Highly Specific and Highly Sensitive Aptamer-Gold Nanoparticle Based NIR Contrast

描述(申请人提供)：荧光团一直是光学成像不可或缺的信号媒介。如果这种荧光造影剂是针对目标分子的，并且具有更高的灵敏度，那么它可能会更有效。金纳米颗粒(GNPs)表面的电磁场(等离子激元)可以通过人工猝灭/增强荧光来帮助这种造影剂具有这些性质。对于分子靶向特定的生物分子，适体被发现与传统的靶向剂单抗一样好。适配子还具有其他有益的性质，如生产更容易和更便宜，更小的尺寸，几乎没有免疫原性等。通过结合荧光团、GNP和适配子的有益性质，我们建议开发一种分子信标状纳米实体，用于早期和准确地检测侵袭性乳腺癌，具有高灵敏度和特异性；也用于同时监测治疗方案的疗效。即将开发的纳米荧光剂是一种表达基质金属蛋白酶-14的乳腺癌类型，被认为是乳腺癌侵袭性的标志物。该实体的成分包括具有两个胸腺嘧啶核苷酸序列的乳腺癌靶向适配子AS1411。序列的一端与GNP结合，另一序列的一端与Cypate结合，Cypate是ICG的类似物(FDA批准的近红外荧光团)。链的长度大约是产生GNP最大荧光的距离的一半。链通过基质金属蛋白酶-14的底物连接，底物很小，因此使GNP和Cypate非常接近，迫使完全猝灭。最初，由于GNP的强等离激元场猝灭了荧光，该实体发出的荧光很少。当它应用于具有侵袭性的乳腺癌患者体内时，它通过AS1411受体的靶点特异性地传递到乳腺癌。然后呢，它 会暴露于肿瘤细胞壁上/内的基质金属蛋白酶-14。当底物被基质金属蛋白酶-14切割时，Cypate从GNP移动到由链长指定的距离，这导致在激发光作用下荧光发射高度增强。我们将这个实体命名为AptaPET(适体-纳米粒子等离子能量转移)，它既可以检测癌症，又可以为我们的系统提供关于癌症性质的信息，即癌症的侵袭性。通过选择具有治疗特性的特殊适体AS1411，该实体也可以用于检测和治疗。此外，在不进一步修饰的情况下，AptaPET可以作为一种良好的X射线/CT造影剂。同样的概念也可以用于其他疾病和其他类型癌症的检测、诊断和预后。

DESCRIPTION (provided by applicant): Fluorophores have been indispensable signal mediators for optical imaging. This fluorescent contrast agent can be more effective if it is specific for the target molecule and with higher sensitivity. The surface electro-magnetic (plasmon) field on the gold nanoparticles (GNPs) can be helpful for this contrast agent to have these properties by allowing artificial quenching/enhancing fluorescence. For molecular targeting particular biomolecules, aptamers are found to be as good as monoclonal antibodies, the traditional targeting agent. Aptamers also have additional beneficial properties, such as easier and cheaper production, smaller size, little immunogenicity, etc. By combining beneficial properties of fluorophores, GNPs, and aptamers, we propose to develop a molecular beacon-like nano-entity for early and accurate detection of aggressive breast cancer, with high sensitivity and specificity; and also for concurrently monitoring the efficacy of the therapeutic regimen. The nano-fluorescent agent to be developed is a breast cancer type expressing MMP-14, which is known as a marker for the breast cancer with aggressive nature. The components of this entity include a breast cancer targeting aptamer AS1411 with two Tymidine nucleotide sequences. One end of a sequence is bound to a GNP and one end of the other sequence is conjugated to Cypate, analogue of ICG (FDA approved near infrared fluorophore). The length of the chain is to be approximately a half of the distance producing the maximum fluorescence of Cypate by a GNP. The chains are connected via the substrate of MMP-14, which is small and, therefore, places the GNP and Cypate very close, forcing complete quenching. Initially, this entity emits little fluorescence because the strong plasmon field of GNP quenches the fluorescence. When it is applied to human body with breast cancer with aggressive nature, it is delivered to the breast cancer specifically via the targeting site of the AS1411 receptor. Then, it will be exposed to the MMP-14 on/in the tumor cell wall. As the substrate gets cleaved by the MMP-14, Cypate moves away from the GNP to the distance specified by the chain length, which results in highly enhanced fluorescence emission upon excitation light application. We have named this entity AptaPET (Aptamer-nanoparticle Plasmon Energy Transfer) and It allows both cancer detection and provides the information on nature of the cancer, for our system, the aggressiveness of the cancer. By selecting a special aptamer AS1411 with therapeutic properties, this entity can also be used both for detection and treatment. In addition, with no further modification, the AptaPET can be a good contrast agent for X-ray/CT. The same concept can be used for detection, diagnosis, and prognosis of other diseases as well as of other types of cancer.