Sulfatase Activated Fluorescent Probes for In Vivo Diagnostic Imaging of Cancer

描述（由申请人提供）：硫酸酯酶催化硫酸酯水解，改变许多在发育、信号传导、降解、发病机制和疾病中具有关键功能的底物。最近表征的硫酸酯酶HSulf-1和HSulf-2的表达水平在某些癌症（例如乳腺癌、胰腺癌、结肠癌）中增加，使得可以通过定量和可视化肿瘤细胞中的硫酸酯酶活性来实现癌症的早期诊断。敏感和选择性的检测需要设计高亲和力的细胞外硫酸酯酶HSulf-1和HSulf-2的荧光探针。该建议描述了荧光团的设计，其在通过细胞外硫酸酯酶水解后应经历量子产率或发射的显著改变。由于一种酶可以水解许多探针，硫酸酯酶激活的荧光信号将被放大，从而能够灵敏地检测异常的癌症相关活性。具有红色或近红外发射的荧光团将被靶向以实现更深的组织穿透并减少组织自发荧光。不同的可水解的荧光团支架将进行评估，包括硫酸化二甲基吖啶酮，试卤灵，二萘嵌苯二酮，和发光香豆素镧系元素共轭物。将在体外、癌细胞系和小鼠肿瘤模型中测定每种探针的光化学和动力学性质。虽然已知硫酸酯酶可以水解多种芳基硫酸酯，但对染料支架的轻微修饰（例如，将检查结合亲和力或荧光性质。探针-酶亲和力将通过与通过筛选DMA适体、肽或类肽文库鉴定的阴离子分子缀合而进一步增强。总之，硫酸酯酶激活的探针将能够灵敏和选择性地对活动物体内的硫酸酯酶活性进行荧光成像，用于癌症的诊断。* 硫酸酯酶在某些癌症中水平升高，包括乳腺癌、胰腺癌和结肠癌。通过它们的活性，这些酶可以通过影响细胞表面的相互作用来控制癌症的严重程度。所描述的探针被细胞外硫酸酯酶激活以在癌细胞处给出不同的荧光信号。这些探针旨在实现早期和准确的癌症诊断。

DESCRIPTION (provided by applicant): Sulfatase enzymes, which catalyze the hydrolysis of sulfate esters, alter many substrates which have key functions in development, signaling, degradation, pathogenesis, and disease. The expression levels of the recently characterized sulfatases HSulf-1 and HSulf-2 are increased in certain cancers (e.g. breast, pancreas, colon), making early diagnosis of cancer achievable by the quantification and visualization of sulfatase activity in tumor cells. Sensitive and selective detection necessitates the design of high affinity fluorescent probes for the extracellular sulfatases HSulf-1 and HSulf-2. This proposal describes the design of fluorophores which should undergo significant alterations in quantum yield or emission after hydrolysis by extracellular sulfatases. Because one enzyme can hydrolyze many probes, the sulfatase-activated fluorescent signal will be amplified, enabling sensitive detection of abnormal, cancer-associated activity. Fluorophores with red or near-infrared emission will be targeted to enable deeper tissue penetration and to reduce tissue autofluorescence. Distinct hydrolyzable fluorophore scaffolds will be evaluated, including sulfated dimethylacridinones, resorufins, perylene diones, and luminogenic coumarin-lanthanide conjugates. The photochemical and kinetic properties will be determined for each probe in vitro, in cancer cell lines, and in mouse tumor models. Although it is known that sulfatases can hydrolyze diverse aryl sulfates, slight modifications to the dye scaffolds (e.g., fluorination) will be examined to improve the binding affinity or fluorescent properties. The probe-enzyme affinity will be further enhanced by conjugation to anionic molecules identified by screening libraries of DMA aptamers, peptides, or peptoids. In summary, the sulfatase activated probes will enable sensitive and selective fluorescence imaging of sulfatase activity inside living animals for the diagnosis of cancer. ***Sulfatases have elevated levels in certain cancers, including breast, pancreatic, and colon cancer. Through their activity, these enzymes may be controlling the severity of cancers by influencing interactions at the cell surface. The described probes are activated by extracellular sulfatases to give a distinct fluorescent signal at cancer cells. The probes are designed to enable early and accurate cancer diagnosis.