Key Laboratory of Chemical Biology (Ministry of Education), School of Pharmaceutical Science , Shandong University , Ji'nan 250012 , Shandong , P. R. China.
School of Chemistry and Chemical Engineering , Shandong University , Ji'nan 250100 , Shandong , P. R. China.
ACS Appl Mater Interfaces. 2019 Oct 2;11(39):35548-35555. doi: 10.1021/acsami.9b11340. Epub 2019 Sep 17.
Overexpression of adenosine 5'-triphosphate-binding cassette transporters is one of the primary causes of drug resistance in cancer. Downregulating the expression of these transporters by inhibiting the mRNA translation process is an effective approach to cope up with this situation. Herein, multifunctional molecular beacons (MBs)-modified gold nanoparticle (AuNP) as a nanocarrier (MBs-AuNP) is developed for synergistic inhibition and in situ imaging of drug-resistant-related mRNAs in living cells. MBs-AuNP is composed of (i) triple specially designed molecular beacons modified on the surface of AuNP, for binding drug-resistant-related mRNAs, loading doxorubicin (Dox), and reporting the fluorescence signal, and (ii) AuNP, for loading MBs, introducing them into cells, and quenching their fluorescence. After uptake by cells, MBs-AuNP will hybridize with three different drug-resistant-related mRNAs (MDR1 mRNA, MRP1 mRNA, and BCRP mRNA), respectively, which could inhibit their translation to decrease efflux protein expression and lead to AuNP-quenched fluorescence recovery for in situ imaging. Real-time quantitative-polymerase chain reaction and western blot results showed that drug-resistant-related mRNAs and efflux proteins expression both decreased. Dox-loaded MBs-AuNP exhibited higher suppression efficacy compared to that of free Dox against HepG2/ADR (0.35 vs 1.06 μM of IC) and MCF-7/ADR (2.78 vs >5 μM of IC). Direct observation of intracellular hybridization events and differentiation of drug-resistant cancer cells or non-drug-resistant cancer cells could be accomplished through fluorescence imaging analysis. This nanocarrier is capable of downregulating the expression of multiple efflux proteins by gene silencing, allows in situ monitoring of silencing events, and thus provides a powerful strategy to cope up with drug resistance at the gene level.
多腺苷酸结合盒转运蛋白的过度表达是癌症药物耐药的主要原因之一。通过抑制 mRNA 翻译过程来下调这些转运蛋白的表达是应对这种情况的有效方法。在此，开发了多功能分子信标（MBs）修饰的金纳米粒子（AuNP）作为纳米载体（MBs-AuNP），用于协同抑制和原位成像活细胞中耐药相关的 mRNA。MBs-AuNP 由（i）三重专门设计的分子信标修饰在 AuNP 表面，用于结合耐药相关的 mRNA、装载阿霉素（Dox）和报告荧光信号，和（ii）AuNP，用于装载 MBs、将其引入细胞并猝灭其荧光。被细胞摄取后，MBs-AuNP 将分别与三种不同的耐药相关的 mRNA（MDR1 mRNA、MRP1 mRNA 和 BCRP mRNA）杂交，从而抑制其翻译以减少外排蛋白表达，并导致 AuNP 猝灭的荧光恢复进行原位成像。实时定量聚合酶链反应和 Western blot 结果表明，耐药相关的 mRNA 和外排蛋白的表达均降低。与游离 Dox 相比，载有 Dox 的 MBs-AuNP 对 HepG2/ADR（0.35 vs 1.06 μM 的 IC）和 MCF-7/ADR（2.78 vs >5 μM 的 IC）具有更高的抑制效果。通过荧光成像分析可以直接观察到细胞内杂交事件以及耐药癌细胞或非耐药癌细胞的分化。这种纳米载体能够通过基因沉默下调多种外排蛋白的表达，允许对沉默事件进行原位监测，从而为应对基因水平的耐药性提供了一种强大的策略。
