Near-Infrared Fluorescent Choline Kinase Inhibitors for Cancer Imaging and Therap

描述（由申请人提供）：本研究的目的是使用能够破坏癌细胞中脂质代谢的近红外（NIR）荧光胆碱类似物对乳腺肿瘤中胆碱激酶（ChoK）的表达进行成像。乳腺癌是影响女性的最常见的癌症，虽然许多乳腺癌患者对目前的护理标准反应良好，但仍有一些亚型的疾病对现有疗法有抗药性。虽然 早期诊断改善了这些患者的预后，但缺乏非侵入性方法来鉴定与这些恶性肿瘤相关的生物标志物。ChoK是一种癌基因，在39%的乳腺癌中过表达，是肿瘤侵袭性的标志物。ChoK表达增加与乳腺癌肿瘤分级相关，表达与雌激素受体状态成反比。已经生产了许多专门针对ChoK的抗癌药物。其中一种抑制剂，TCD-717，目前正在进行I期临床试验，用于治疗晚期实体瘤。检测乳腺癌中该癌基因表达的方法的开发对于乳腺癌状态的非侵入性分期和适合于ChoK靶向治疗的患者的鉴定具有高度意义。近红外（NIR）光学成像是一种相对便宜的技术，它采用灵敏的示踪剂，而不使用电离辐射，以报告体内特定的分子相互作用。在动物模型中已知的最有效的ChoK抑制剂MN 58 b与用于NIR光学成像的主要类别的化合物花青染料具有密切的结构相似性。该项目旨在利用这种相似性来设计NIR荧光胆碱类似物，其在与MN 58 b相当的浓度下有效抑制ChoK。我们的原型化合物JAS 239具有许多有利的特性：它独立于胆碱转运蛋白进入癌细胞，定位于ChoK正常表达的细胞质，在动物肿瘤的非缺氧区域积累，并通过过量胆碱的补救有效逆转。拟议的工作将包括综合微调原型NIR ChoK类似物的结构，以优化体外抑制特性和体内生物分布。使用14 C-胆碱放射性示踪、活细胞和细胞提取物的NMR、荧光测定法和共聚焦显微镜，将表征荧光胆碱类似物的效力、NIR荧光和结合选择性。然后将在具有不同ChoK表达的鼠乳腺癌模型中评估最佳候选物。将使用每种荧光团的痕量水平探索诊断潜力，由于ChoK结合，预计最特异的探针将在最具侵袭性的肿瘤中积累。然后将配制更高剂量的方案，并将肿瘤生长延迟和动物存活测量与MN 58 b进行比较。

DESCRIPTION (provided by applicant): The aim of this research is to image the expression of choline kinase (ChoK) in breast tumors using near-infrared (NIR) fluorescent choline analogs capable of disrupting lipid metabolism in cancer cells. Breast cancer is the most prevalent cancer affecting women, and while many breast cancer patients respond well to the current standard of care, there are still subtypes of the disease resistant to existing therapies. Although early diagnosis improves the prognosis for these patients, there is a shortage of non-invasive methods to identify biomarkers associated with these malignant tumors. ChoK is an oncogene that is overexpressed in 39% of breast cancers, and is a marker of tumor aggressiveness. Increased ChoK expression is correlated with breast cancer tumor grade and the expression is inversely proportional to estrogen receptor status. A number of anticancer drugs specifically targeting ChoK have been produced. One of these inhibitors, TCD-717, is now being tested in Phase I Clinical Trials for treatment of advanced solid tumors. The development of methods to detect the expression of this oncogene in breast cancer would be highly significant for non-invasive staging of breast cancer status and for the identification of patients suited for ChoK-targeted therapy. Near-infrared (NIR) optical imaging is a relatively inexpensive technique that employs sensitive tracers, without the use of ionizing radiation, to report specific molecular interactions in vivo. The most effective known inhibitor of ChoK in animal models, MN58b, bears close structural similarity to the cyanine dyes, the major class of compounds that are used for NIR optical imaging. This project seeks to exploit this similarity to design NIR-fluorescent choline analogs that effectively inhibit ChoK at concentrations comparable to MN58b. Our prototype compound, JAS239, has a number of favorable characteristics: it enters cancer cells independently of the choline transporters, localizes to the cytoplasm where ChoK is normally expressed, accumulates in non-hypoxic regions of animal tumors, and is effectively reversed by salvage with excess choline. The proposed work will include synthetically fine-tuning the structure of the prototype NIR ChoK analog to optimize the inhibitory properties in vitro and bio-distribution in vivo. Using 14C-choline radio-tracing, NMR of live cells and cell extracts, fluorimetry, and confocal microscopy, the fluorescent choline analogs will be characterized for potency, NIR-fluorescence, and binding selectivity. The top candidates will then be assessed in murine breast cancer models with varying ChoK expression. The diagnostic potential will be explored using trace levels of each fluorophore, with the most specific probes expected to accumulate in the most aggressive tumors due to ChoK-binding. Higher-dose regimens will then be formulated, and tumor growth delay and animal survival measurements compared to MN58b.