SELECTIVELY IMAGING CANCER STEM CELLS

描述(申请人提供)：越来越多的证据表明，肿瘤生长是由肿瘤干细胞(CSCs)驱动的。这些细胞被认为是肿瘤生长的驱动力，因为它们对治疗具有抵抗力，并且能够自我更新。在发现CSCs之前，肿瘤进展的克隆模型得到了大多数支持。该模型假设，肿瘤内的每个细胞都具有自我更新的能力，因此，如果在治疗过程中没有被杀死，可以用对治疗具有抵抗力的亚克隆重新填充肿瘤。CSC模型认为，肿瘤由一系列细胞组成。肿瘤内的CSC是能够自我再生并共同产生非CSC后代的细胞，非CSC后代构成了剩余肿瘤体积的很大一部分。由于CSCs是一组肿瘤细胞中的一个亚群，它们被标记了两个不同的分子，这些分子附着在细胞的外部。我们提出了一种基于抗体的成像系统，这些抗体对我们将要开发的两个标记具有高亲和力。我们还将为一种小报告分子开发第三种抗体，该抗体将被一种染料标记。这种抗体将被一分为二，每一半将与两个细胞表面标志物的抗体融合。当细胞表面标记被抗体结合时，这两个部分将紧密结合在一起，形成一个小报告分子的结合口袋。我们将通过在显微镜下观察人类肿瘤细胞的过程来可视化小报告分子是否结合，从而证明这个系统是有效的。小报告分子将携带一种染料，使其能够可视化。下文目标1-3概述了这一进程。目的1.制备和检测抗细胞表面标志物和小报告分子的抗体。目的2将半抗体与全抗体进行融合，再进行检测，表明抗体的结合能力没有受到影响。目的3将融合抗体和染料标记的小报告分子加入到人癌细胞中，在显微镜下观察这一过程，以确定该系统是否能够工作。如果我们能够使这种成像系统工作，那么它就可以在人类癌症的动物模型上进行测试。如果成功，它也可以用于人类观察治疗过程中和治疗后的CSCs水平。当抗CSC疗法被开发出来时，这将特别有用。

DESCRIPTION (provided by applicant): Evidence is mounting that tumor growth is driven by cancer stem cells (CSCs).These cells are believed to act as the driving force for tumor growth because they are resistant to therapy and are able to self renew. Prior to the discovery of CSCs, the clonal model of tumor progression held the majority of support. This model posits that every cell within a tumor has the capacity for self-renewal and therefore, if not killed during therapy, can repopulate the tumor with subclones resistance to the therapy. The CSC model maintains that a tumor consists of a hierarchy of cells. The CSCs within the tumor are the cells that are able to regenerate themselves and co-produce non-CSC progeny which make up a good deal of the remaining tumor bulk. Because CSCs are a sub population within a group of tumor cells, they are marked with two different molecules that are attached to the outside of the cell. We propose an imaging system based on antibodies that have high affinity for two of these markers which we will develop. We will also develop a third antibody for a small reporting molecule that will be labeled with a dye. This antibody will be divided in half and each half will be fused to the antibodies for the two cell surface markers. When the cell surface markers are bound by the antibodies the two halves will be brought in close proximity and form a binding pocket for the small reporting molecule. We will prove this system works by visualizing whether the small reporting molecule binds by viewing the process on human tumor cells in a microscope. The small reporting molecule will bear a dye that will enable its visualization. The process is outlined below in Aims 1-3. Aim 1. Prepare and test antibodies to the cell surface markers and the small reporting molecule. Aim 2 Fuse the half-antibodies to the whole antibodies and retest to show that binding ability has not been hampered. Aim 3 Add the fused antibodies and dye-labeled small reporting molecule to the human cancer cells and view the process under the microscope to determine whether the system can work. If we are able to make this imaging system work, it can then be tested in animal models of human cancer. If successful, it also be used in humans to view the levels of CSCs during and after therapy. This will be particularly useful when anti-CSC therapies are developed.