（钆-二乙醇三胺五乙酸）n-聚二氨基丙酰-肽核酸-D（半胱氨酸-丝氨酸-赖氨酸-半胱氨酸）磁共振造影剂的设计

Design of (Gd-DO3A)n-polydiamidopropanoyl-peptide nucleic acid-D(Cys-Ser-Lys-Cys) magnetic resonance contrast agents.

作者信息

Amirkhanov Nariman V, Dimitrov Ivan, Opitz Armin W, Zhang Kaijun, Lackey John P, Cardi Christopher A, Lai Song, Wagner Norman J, Thakur Mathew L, Wickstrom Eric

机构信息

Laboratory of Nucleic Acids Chemistry, Institute of Chemical Biology and Fundamental Medicine, Philadelphia, PA 19107, USA.

出版信息

Biopolymers. 2008 Dec;89(12):1061-76. doi: 10.1002/bip.21059.

We hypothesized that chelating Gd(III) to 1,4,7-tris(carboxymethylaza)cyclododecane-10-azaacetylamide (DO3A) on peptide nucleic acid (PNA) hybridization probes would provide a magnetic resonance genetic imaging agent capable of hybridization to a specific mRNA. Because of the low sensitivity of Gd(III) as an magnetic resonance imaging (MRI) contrast agent, a single Gd-DO3A complex per PNA hybridization agent could not provide enough contrast for detection of cancer gene mRNAs, even at thousands of mRNA copies per cell. To increase the Gd(III) shift intensity of MRI genetic imaging agents, we extended a novel DO3An-polydiamidopropanoyl (PDAPm) dendrimer, up to n = 16, from the N-terminus of KRAS PNA hybridization agents by solid phase synthesis. A C-terminal D(Cys-Ser-Lys-Cys) cyclized peptide analog of insulin-like growth factor 1 (IGF1) was included to enable receptor-mediated cellular uptake. Molecular dynamic simulation of the (Gd-DO3A-AEEA)16-PDAP4-AEEA2-KRAS PNA-AEEA-D(Cys-Ser-Lys-Cys) genetic imaging nanoparticles in explicit water yielded a pair correlation function similar to that of PAMAM dendrimers, and a predicted structure in which the PDAP dendron did not sequester the PNA. Thermal melting measurements indicated that the size of the PDAP dendron included in the (DO3A-AEEA)n-PDAPm-AEEA2-KRAS PNA-AEEA-D(Cys-Ser-Lys-Cys) probes (up to 16 Gd(III) cations per PNA) did not depress the melting temperatures (Tm) of the complementary PNA/RNA hybrid duplexes. The Gd(III) dendrimer PNA genetic imaging agents in phantom solutions displayed significantly greater T1 relaxivity per probe (r1 = 30.64 +/- 2.68 mM(-1) s(-1) for n = 2, r1 = 153.84 +/- 11.28 mM(-1) s(-1) for n = 8) than Gd-DTPA (r1 = 10.35 +/- 0.37 mM(-1) s(-1)), but less than that of (Gd-DO3A)32-PAMAM dendrimer (r1 = 771.84 +/- 20.48 mM(-1) s(-1)) (P < 0.05). Higher generations of PDAP dendrimers with 32 or more Gd-DO3A residues attached to PNA-D(Cys-Ser-Lys-Cys) genetic imaging agents might provide greater contrast for more sensitive detection.

我们推测，将钆（III）螯合到肽核酸（PNA）杂交探针上的1,4,7 - 三（羧甲基氮杂）环十二烷 - 10 - 氮杂乙酰酰胺（DO3A）上，会产生一种能够与特定信使核糖核酸（mRNA）杂交的磁共振基因成像剂。由于钆（III）作为磁共振成像（MRI）造影剂的灵敏度较低，即使每个细胞中有数千个mRNA拷贝，每个PNA杂交剂中的单个钆 - DO3A复合物也无法提供足够的对比度来检测癌症基因mRNA。为了提高MRI基因成像剂的钆（III）位移强度，我们通过固相合成从KRAS PNA杂交剂的N端延伸出一种新型的DO3An - 聚二氨基丙酰基（PDAPm）树枝状聚合物，直至n = 16。包含胰岛素样生长因子1（IGF1）的C端D（半胱氨酸 - 丝氨酸 - 赖氨酸 - 半胱氨酸）环化肽类似物，以实现受体介导的细胞摄取。在显式水中对（钆 - DO3A - AEEA）16 - PDAP4 - AEEA2 - KRAS PNA - AEEA - D（半胱氨酸 - 丝氨酸 - 赖氨酸 - 半胱氨酸）基因成像纳米颗粒进行分子动力学模拟，得到了与聚酰胺 - 胺（PAMAM）树枝状聚合物相似的对关联函数，以及预测的结构，其中PDAP树枝状结构未隔离PNA。热熔测量表明，（DO3A - AEEA）n - PDAPm - AEEA2 - KRAS PNA - AEEA - D（半胱氨酸 - 丝氨酸 - 赖氨酸 - 半胱氨酸）探针中包含的PDAP树枝状结构的大小（每个PNA最多16个钆（III）阳离子）并未降低互补PNA / RNA杂交双链体的解链温度（Tm）。模型溶液中的钆（III）树枝状PNA基因成像剂每个探针显示出明显更高的纵向弛豫率（对于n = 2，r1 = 30.64±2.68 mM-1 s-1；对于n = 8，r1 = 153.84±11.28 mM-1 s-1），高于钆 - 二乙三胺五乙酸（DTPA）（r1 = 10.35±0.37 mM-1 s-1），但低于（钆 - DO3A）32 - PAMAM树枝状聚合物（r1 = 771.84±20.48 mM-1 s-1）（P < 0.05）。连接到PNA - D（半胱氨酸 - 丝氨酸 - 赖氨酸 - 半胱氨酸）基因成像剂上的具有32个或更多钆 - DO3A残基的更高代数的PDAP树枝状聚合物可能提供更高的对比度以实现更灵敏的检测。