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

描述(申请人提供)：这项研究的目的是使用能够扰乱癌细胞脂质代谢的近红外(NIR)荧光胆碱类似物来成像胆碱激酶(CHOK)在乳腺肿瘤中的表达。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，但仍有一些疾病亚型对现有的治疗方法产生抗药性。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识别与这些恶性肿瘤相关的生物标志物。Chok是一种癌基因，在39%的乳腺癌中过度表达，是肿瘤侵袭性的标志。CHOK表达增加与乳腺癌肿瘤分级相关，与雌激素受体状态成反比。一些专门针对Chok的抗癌药物已经生产出来。其中一种抑制剂TCD-717目前正在进行I期临床试验，用于治疗晚期实体肿瘤。该癌基因在乳腺癌中表达的检测方法的发展对乳腺癌状态的非侵袭性分期和适合CHOK靶向治疗的患者的识别具有重要意义。近红外(NIR)光学成像是一种相对便宜的技术，它使用敏感的示踪剂，而不使用电离辐射，来报告体内特定的分子相互作用。在动物模型中，已知最有效的ChOK抑制剂MN58b与用于近红外光学成像的主要化合物菁染料在结构上具有密切的相似性。这个项目试图利用这种相似性来设计近红外荧光胆碱类似物，在与MN58b相当的浓度下有效地抑制Chok。我们的原型化合物JAS239具有许多有利的特征：它独立于胆碱转运蛋白进入癌细胞，定位于正常表达Chok的细胞质，在动物肿瘤的非缺氧区蓄积，并可通过挽救过量胆碱有效地逆转。拟议的工作将包括对原型近红外CHOK类似物的结构进行综合微调，以优化其体外抑制性能和体内生物分布。利用~(14)C-胆碱放射性示踪、活细胞和细胞提取物的核磁共振、荧光测定法和共聚焦显微镜，荧光胆碱类似物的效力、近红外荧光和结合选择性将被表征。然后将在具有不同Chok表达的小鼠乳腺癌模型中对排名靠前的候选药物进行评估。将使用每个荧光团的痕量水平来探索诊断潜力，由于Chok结合，最特异的探针预计会积聚在最具侵袭性的肿瘤中。然后将制定更高剂量的方案，与MN58b相比，肿瘤生长延迟和动物存活测量。

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.