Cheng Jianjun, Zeidan Ryan, Mishra Swaroop, Liu Aijie, Pun Suzie H, Kulkarni Rajan P, Jensen Gregory S, Bellocq Nathalie C, Davis Mark E
Insert Therapeutics, 2585 Nina Street, Pasadena, CA 91107, USA.
J Med Chem. 2006 Nov 2;49(22):6522-31. doi: 10.1021/jm060736s.
To understand how chloroquine (CQ) enhances transgene expression in polycation-based, nonviral gene delivery systems, a number of CQ analogues with variations in the aliphatic amino side chain or in the aromatic ring are synthesized and investigated. Our studies indicate that the aliphatic amino moiety of CQ is essential to provide increased gene expression. Further, the enhancements are more dramatically affected by changes to the aromatic ring and are positively correlated to the strength of intercalation between DNA and the CQ analogues. Quinacrine (QC), a CQ analogue with a fused acridinyl structure that can strongly intercalate DNA, enhances transfection similarly to CQ at a concentration 10 times lower, while N(4)-(4-pyridinyl)-N(1),N(1)-diethyl-1,4-pentanediamine (CP), a CQ analogue that has a weakly intercalating pyridinyl ring, shows no effect on gene expression. Subtle change on the 7-substituent of the chloroquine aromatic structure can also greatly affect the ability of the CQ analogues to enhance transgene expression. Transfection in the presence of N(4)-(7-trifluoromethyl-4-quinolinyl)-N(1),N(1)-diethyl-1,4-pentanediamin e (CQ7a) shows expression efficiency 10 times higher than in the presence of CQ at same concentration, while transfection in the presence of N(4)-(4-quinolinyl)-N(1),N(1)-diethyl-1,4-pentanediamine (CQ7b) does not reveal any enhancing effects on expression. Through a number of comparative studies with CQ and its analogues, we conclude that there are at least three mechanistic features of CQ that lead to the enhancement in gene expression: (i) pH buffering in endocytic vesicles, (ii) displacement of polycations from the nucleic acids in polyplexes, and (iii) alteration of the biophysical properties of the released nucleic acid.
为了解氯喹（CQ）如何增强基于聚阳离子的非病毒基因递送系统中的转基因表达，合成并研究了一些脂肪族氨基侧链或芳香环有变化的CQ类似物。我们的研究表明，CQ的脂肪族氨基部分对于提高基因表达至关重要。此外，芳香环的变化对增强效果影响更为显著，并且与DNA和CQ类似物之间的嵌入强度呈正相关。喹吖因（QC）是一种具有稠合吖啶基结构、能强烈嵌入DNA的CQ类似物，在浓度低10倍时与CQ类似地增强转染，而N(4)-(4-吡啶基)-N(1),N(1)-二乙基-1,4-戊二胺（CP）是一种具有弱嵌入吡啶环的CQ类似物，对基因表达无影响。氯喹芳香结构7位取代基的细微变化也会极大地影响CQ类似物增强转基因表达的能力。在N(4)-(7-三氟甲基-4-喹啉基)-N(1),N(1)-二乙基-1,4-戊二胺（CQ7a）存在下转染显示，在相同浓度时表达效率比在CQ存在下高10倍，而在N(4)-(4-喹啉基)-N(1),N(1)-二乙基-1,4-戊二胺（CQ7b）存在下转染未显示对表达有任何增强作用。通过对CQ及其类似物的一系列比较研究，我们得出结论，CQ导致基因表达增强至少有三个机制特征：（i）内吞小泡中的pH缓冲，（ii）从多聚体中的核酸上取代聚阳离子，以及（iii）释放的核酸生物物理性质的改变。
