DESIGN OF LIGANDS SELECTIVE FOR THE DAG SUPERFAMILY

DAG超家族选择性调节剂的发现， 细胞中的phorbel酯受体，目前包括PKC， chimaerins和unc-13对于确定 单个蛋白质在生理过程中的作用通知发现 DAG超家族的小分子调节剂在很大程度上是未满足的 药理需要。该提案的主要目的是建立 特异性DAG家族成员的新型非肿瘤促进配体。 目前，我们知道配体如何结合到DAG位点，并且我们知道如何结合 配体可以被修饰以实现DAG之间的选择性 超家族成员发现了改善的DAG位点调节剂 选择性不仅有助于阐明 生物过程中的单个蛋白质，但最终应该 促进发现治疗疾病的新疗法 如癌症和阿尔茨海默病。苔藓抑素提供了一种 DAG超家族调制器的引人注目的例子， 临床上用于治疗癌症。在这项研究的过程中， 我们会致力达致以下目标： .1. ILV与其受体相互作用的分子模拟研究 苯并内酰胺模拟物，以及PKC CRD内的选定合成靶标， 激活剂结合结构域，以及DAG的结合位点， 将进行嵌合体和unc-13。这些研究将指导 设计和选择可能表现出DAG位点的新合成靶标 选择性将开展构效关系研究 为了揭示对结合亲和力和 选择性，从而指导合成新结构的设计。 .2.另外的苯并内酰胺类似物的合成 可以赋予增强的同工酶选择性的取代基将是 使用我们已经开发的化学方法进行。 同时，我们也将探索新型火元环的活性 ILV的类似物[2-吡咯烷酮];初步实验揭示了这些 化合物以良好的亲和力与PKCalpha结合。 .3.为了确定新合成的化合物的DAG家族选择性， 材料，体外结合研究将使用 相关的重组PKC同工酶，包括突变体以及 嵌合体和UNC-13。为了评估改变的影响， 配体的疏水侧链对其促肿瘤活性的影响，研究 将进行皮肤增生检查。

The discovery of selective modulators of the DAG superfamily, the major phorbel ester receptors in cells, which presently includes PKC, the chimaerins, and unc-13 is crucial to ascertaining the role of the individual proteins in physiological processes. The discovery of such small molecule modulators of the DAG superfamily is largely an unmet pharmacological need. The major objective of this proposal is to create novel, non-tumor promoting ligands for specific DAG family members. Presently, we know how ligands bind to the DAG site, and we know how ligands might be modified to achieve selectivity among the DAG superfamily members. The discovery of DAG-site modulators of improve selectivity will not only aid the elucidation of the roles of the individual proteins in biological processes, but should eventually foster the discovery of novel therapeutics for the treatment of disease states such as cancer and Alzheimer's disease. Bryostatin provides a compelling example of a DAG superfamily modulator that has found use clinically in the treatment of cancer. During the course of this study, we will undertake the following aims: .1. Molecular modeling studies of the interactions of ILV, its benzolactam mimic, and selected synthetic targets within the PKC CRDs, the activator-binding domains, as well as the DAG binding site of the chimaerins and unc-13 will be conducted. These studies will guide the design and selection of new synthetic targets likely to exhibit DAG-site selectivity. Structure-activity relationship studies will be carried out to uncover structural features important to both binding affinity and selectivity, thus guiding the design of novel structures of synthesis. .2. The synthesis of additional benzolactam analogues bearing substituents that may confer enhanced isozyme selectivity will be carried out using chemical methods already developed by us. Simultaneously, we will explore the activity of novel fire-membered ring analogues [2-pyrrolidones] of ILV; preliminary experiments reveal these compounds to bind with good affinity to PKCalpha. .3. To determine the DAG family selectivity of the newly synthesized materials, in vitro binding studies will be carried out using the relevant recombinant PKC isozymes including mutants as well as the chimaerins and unc-13. To assess the effect of alterations in the ligand's hydrophobic side chain on its tumor promoting activity, studies of skin hyperplasia will be conducted.