Chen Shih-Hsien, Kuo Yu-Ting, Singh Gyan, Cheng Tian-Lu, Su Yu-Zheng, Wang Tzu-Pin, Chiu Yen-Yu, Lai Jui-Jen, Chang Chih-Ching, Jaw Twei-Shiun, Tzou Shey-Cherng, Liu Gin-Chung, Wang Yun-Ming
Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University , 100 Shih-Chuan first Road, Kaohsiung 807, Taiwan.
Inorg Chem. 2012 Nov 19;51(22):12426-35. doi: 10.1021/ic301827p. Epub 2012 Nov 1.
β-Glucuronidase is a key lysosomal enzyme and is often overexpressed in necrotic tumor masses. We report here the synthesis of a pro receptor-induced magnetization enhancement (pro-RIME) magnetic resonance imaging (MRI) contrast agent ([Gd(DOTA-FPβGu)]) for molecular imaging of β-glucuronidase activity in tumor tissues. The contrast agent consists of two parts, a gadolinium complex and a β-glucuronidase substrate (β-d-glucopyranuronic acid). The binding association constant (KA) of [Gd(DOTA-FPβGu)] is 7.42 × 10(2), which is significantly lower than that of a commercially available MS-325 (KA = 3.0 × 10(4)) RIME contrast agent. The low KA value of [Gd(DOTA-FPβGu)] is due to the pendant β-d-glucopyranuronic acid moiety. Therefore, [Gd(DOTA-FPβGu)] can be used for detection of β-glucuronidase through RIME modulation. The detail mechanism of enzymatic activation of [Gd(DOTA-FPβGu)] was elucidated by LC-MS. The kinetics of β-glucuronidase catalyzed hydrolysis of [Eu(DOTA-FPβGu)] at pH 7.4 best fit the Miechalis-Menten kinetic mode with Km = 1.38 mM, kcat = 3.76 × 10(3), and kcat/Km = 2.72 × 10(3) M(-1) s(-1). The low Km value indicates high affinity of β-glucuronidase for [Gd(DOTA-FPβGu)] at physiological pH. Relaxometric studies revealed that T1 relaxivity of [Gd(DOTA-FPβGu)] changes in response to the concentration of β-glucuronidase. Consistent with the relaxometric studies, [Gd(DOTA-FPβGu)] showed significant change in MR image signal in the presence of β-glucuronidase and HSA. In vitro and in vivo MR images demonstrated appreciable differences in signal enhancement in the cell lines and tumor xenografts in accordance to their expression levels of β-glucuronidase.
β-葡萄糖醛酸酶是一种关键的溶酶体酶，在坏死肿瘤块中常过度表达。我们在此报告一种用于肿瘤组织中β-葡萄糖醛酸酶活性分子成像的前体受体诱导磁化增强（pro-RIME）磁共振成像（MRI）造影剂（[Gd(DOTA-FPβGu)]）的合成。该造影剂由两部分组成，钆配合物和β-葡萄糖醛酸酶底物（β-D-吡喃葡萄糖醛酸）。[Gd(DOTA-FPβGu)]的结合缔合常数（KA）为7.42×10²，显著低于市售的MS-325（KA = 3.0×10⁴）RIME造影剂。[Gd(DOTA-FPβGu)]的低KA值归因于侧链β-D-吡喃葡萄糖醛酸部分。因此，[Gd(DOTA-FPβGu)]可用于通过RIME调制检测β-葡萄糖醛酸酶。通过液相色谱-质谱（LC-MS）阐明了[Gd(DOTA-FPβGu)]的酶促激活详细机制。在pH 7.4条件下，β-葡萄糖醛酸酶催化[Eu(DOTA-FPβGu)]水解的动力学最符合米氏动力学模式，Km = 1.38 mM，kcat = 3.76×10³，kcat/Km = 2.72×10³ M⁻¹ s⁻¹。低Km值表明在生理pH下β-葡萄糖醛酸酶对[Gd(DOTA-FPβGu)]具有高亲和力。弛豫研究表明，[Gd(DOTA-FPβGu)]的T1弛豫率随β-葡萄糖醛酸酶浓度而变化。与弛豫研究一致，[Gd(DOTA-FPβGu)]在存在β-葡萄糖醛酸酶和人血清白蛋白（HSA）时，磁共振图像信号有显著变化。体外和体内磁共振图像显示，根据细胞系和肿瘤异种移植中β-葡萄糖醛酸酶的表达水平，信号增强存在明显差异。
