Multimeric Ligands for Targeting Melanoma

描述(由申请人提供)：拟议工作的目标是开发针对转移性黑色素瘤的靶向成像和治疗的异二价物。这些试剂包含两个配体，指导构建物交联两个不同的细胞表面受体，导致结合选择性的显著提高。这比针对单一受体的药物有优势，因为它们不依赖于单一细胞表面蛋白的过度表达。与其他多功能试剂相比，所建议的结构也具有优势，因为它们是通过聚合合成产生的，并且设计得相对较小，具有良好的ADME特性。在第一个支持阶段的工作已经(1)确定并验证了一种可用于靶向转移性黑色素瘤的子集的受体组合，以及(2)证明了合成的异价药物可以使异种受体交联的原理证明。与相应的单价相互作用相比，这些结构结合的亲和力高达50倍。目前的研究将结合这两个方面的进展来开发针对已识别的靶受体对的异二价药物。为了实现这一目标，Aim 1将使用先前开发的G蛋白偶联受体(GPCR)系统来推导预测多价配体作为显像剂的行为的分析解决方案。目的2将开发针对第一期支持中确定的受体组合的二价体：N-甲酰基多肽受体样2型(FPRL2)和1型黑素皮质素受体(MC1R)。在这个目标中，最佳的连接物和配基化学将通过高通量结合分析来确定。二价配体将用荧光团标记，以便通过细胞内和体内成像进行测试。那些具有最佳特性的将被标记为DOTA。DOTA衍生物将用于螯合Eu用于体外荧光，111In用于体内SPECT药代动力学和生物分布的评估。AIM 3将继续对在第一个支持阶段被确定为靶标的另外21个受体进行验证。这些将主要通过包含多个黑色素瘤分级的组织微阵列以及多个正常组织的免疫组织化学来验证。预计这项工作将导致针对大多数转移性黑色素瘤的额外受体组合的验证。在下一个支持阶段结束时，我们将(1)开发更精确的分析方法来预测多价配体作为显像剂的行为；(2)开发对40-50%的转移性黑色素瘤有用的靶向配体；(3)针对这种疾病的剩余分子表型确定额外的3-和4-受体靶向组合。拟议工作的目标是开发平台靶向试剂，用于转移性黑色素瘤的成像和治疗。这些药物是异二价的，因为它们将针对两个不同细胞表面受体的两个配体连接在一起。与针对单一表位的药物相比，这种药物具有优势，因为它不依赖于单一的过度表达的细胞表面蛋白。与其他多功能试剂相比，它们也具有优势，因为它们是通过聚合合成产生的，因此相对较小，具有可接受的ADME特性。

DESCRIPTION (provided by applicant): The goal of the proposed work is to develop heterobivalent agents for targeted delivery of imaging and therapy to metastatic melanoma. These agents contain two ligands that direct the construct to crosslink two different cell surface receptors, resulting in dramatic increases in binding selectivity. This has advantages over agents directed against single receptors in that they do not rely on overexpression of a single cell surface protein. The proposed constructs also have advantages over other multifunctional agents in that they are produced via convergent synthesis and are designed to be relatively small with favorable ADME characteristics. Work during the first period of support has (1) identified and validated a receptor combination that can be used to target a subset of metastatic melanomas, and (2) demonstrated a proof-of-principle that synthetic heterobivalent agents can crosslink heterologous receptors. These constructs bound with up to 50-fold higher affinity compared to corresponding monovalent interactions. Current research will combine these two advances to develop heterobivalent agents against an identified target receptor pair. To achieve this goal, Aim 1 will use a previously developed G-protein coupled receptor (GPCR) system to derive analytical solutions that predict behavior of multivalent ligands as imaging agents. Aim 2 will develop bivalent agents that target a receptor combination identified in the first period of support: N-formyl peptide receptor like, type 2 (FPRL2) and the type 1 melanocortin receptor (MC1R). In this aim, optimum linker and ligand chemistries will be determined with high-throughput binding assays. Bivalent ligands will be labeled with fluorophores for testing by in-cyto and in-vivo imaging. Those with optimum characteristics will be labeled with DOTA. DOTA derivatives will be used to chelate Eu for ex vivo fluorescence and 111In for in vivo SPECT assessment of pharmacokinetics and biodistribution. Aim 3 will continue the validation effort for 21 additional receptors that were identified as targets during the first period of support. These will be validated primarily though immunohistochemistry of tissue microarrays containing multiple melanoma grades as well as multiple normal tissues. It is expected that this work will result in the validation of additional receptor combinations that will target a majority of metastatic melanomas. At the end of this next period of support, we will have (1) developed more precise analytical methods to predict the behavior of multivalent ligands as imaging agents; (2) developed targeting ligands that will be useful against 40-50% of metastatic melanomas and (3) identified additional 3- and 4-receptor target combinations for the remaining molecular phenotypes of this disease.The goal of the proposed work is to develop platform targeting agents for delivery of imaging and therapy to metastatic melanoma. These agents are heterobivalent in that they link together two ligands that are directed against two different cell surface receptors. Such agents have advantages over agents directed against single epitopes by not relying on a single overexpressed cell surface protein. They also have advantages over other multifunctional agents in that they are produced via convergent synthesis and hence are relatively small with acceptable ADME characteristics.