Development of anti-polo-box therapeutic agents

抗polo-box治疗剂的开发

• 批准号: 8157396
• 负责人: Kyung Lee
• 金额: $ 34.28万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8157396
• 关键词: Boxing; Development; Therapeutic Agents

Development of a specific anti-Plk1 therapeutic agent is one of the most compelling missions in the field of anti-cancer therapy. Efforts to generate Plk1-specific inhibitors by targeting the catalytic activity of Plk1 have proven to be difficult due to similarities with the catalytic domains of other structurally related kinases. Here, we propose to develop a new class of mono-specific Plk1 inhibitors by employing a novel approach of targeting the non-catalytic, but functionally essential, PBD of Plk1. To this end, we will take advantage of the crystal structures of the Plk1 PBD in complex with a highly specific ligand, PLHSpT. The first aim of the project will be to develop PLHSpT-derived templates for drug design and subsequent modifications. Since the function of the N-terminal Pro-4 (number indicates the relative position of the residue from pT) residue of PLHSpT can be substituted by hydrophobic moieties and the side chain of the Leu-3 residue is not involved in interactions with surrounding PBD residues, we will first generate N-substituted glycine (Nsg)-containing LHSpT or HSpT peptoidpeptide hybrids and their respective cyclic forms. Then, the Nsg residue of the hybrids will be modified by covalently conjugating them with site-specifically synthesized hydrophobic moieties to fill in the intramolecular cavity present between the compound and the PBD. Since the His-2, Ser-1 and p-Thr residues are critical for the specificity and high affinity binding, these residues will not be modified. Peptide-derived inhibitors are commonly associated with problems in stability, lipophilicity, and transcellular permeability. Hence, the second aim of the project will be to enhance the stability, membrane permeability, and tumor-specific targeting of the above compounds by generating innovative prodrugs. Compounds with a high PBD-binding affinity and specificity will be converted to phosphatase-insensitive, non-hydrolyzable, p-Thr mimetic (Pmab) forms and then further modified to generate Ala- or Val-ester-conjugated phosphonic diamide prodrugs. The latter modification is not only to eliminate the electronegativity of the dianionic phosphonic acid moiety for better transcellular permeation but also to target the compound to the highly active PepT1 transporter for efficient cellular uptake. For the compounds that exhibit anti-Plk1 PBD activities at the cultured cell level, we will investigate whether the addition of a tumor-targeting RGD motif facilitates tumor-specific delivery of the compounds. As the third aim of the project, we will determine the potency and selectivity of the resulting compounds in the inhibition of Plk1-dependent cell proliferation activity in mouse tumor models. Because of the harsh chemical and enzymatic conditions of the gastrointestinal tract, intravenous injection into mouse-tail vein will be the choice of compound administration. Since peptide-derived inhibitors often exhibit physicochemical drawbacks, we will also take complementary approaches to isolate PBD-inhibitory compounds or moieties by screening the NCI natural products extract repository or by carrying out in silico screening. Structural analyses and computer modeling of the isolated small molecule compounds together with the above PLHSpT-derived inhibitors will allow us to perform site-specific replacements and modifications of the latter inhibitors to achieve enhanced in vivo stability and bioavailability. To this end, we will bring the expertise of our collaborators ranging from in vitro high throughput screening and cell-based assays (Dr. James McMahon), in silico screening and computer modeling (Dr. Marc Nicklaus), and X-ray crystallography (Drs. Alex Wlodawer and Michael Yaffe). The ultimate goal of this multifaceted approach is to generate a new class of mono-specific anti-Plk1 therapeutic agents that have potential to treat various cancers in humans.

开发特异性抗Plk 1治疗剂是抗癌治疗领域最引人注目的任务之一。通过靶向Plk 1的催化活性来产生Plk 1特异性抑制剂的努力已被证明是困难的，这是由于与其他结构相关激酶的催化结构域的相似性。在这里，我们建议开发一类新的单特异性Plk 1抑制剂，采用一种新的方法，针对非催化，但功能上必不可少的，PBD的Plk 1。为此，我们将利用Plk 1 PBD与高度特异性配体PLHSpT复合的晶体结构。该项目的第一个目标将是开发用于药物设计和后续修改的PLHSpT衍生模板。由于PLHSpT的N-末端Pro-4（数字表示来自pT的残基的相对位置）残基的功能可以被疏水部分取代，并且Leu-3残基的侧链不参与与周围PBD残基的相互作用，我们将首先产生含N-取代甘氨酸（Nsg）的LHSpT或HSpT类肽杂合体及其各自的环状形式。然后，杂合体的Nsg残基将通过将它们与位点特异性合成的疏水部分共价缀合来修饰，以填充化合物和PBD之间存在的分子内空腔。由于His-2、Ser-1和p-Thr残基对于特异性和高亲和力结合至关重要，因此这些残基将不被修饰。肽衍生的抑制剂通常与稳定性、亲脂性和跨细胞渗透性的问题相关。因此，该项目的第二个目标将是通过产生创新的前药来增强上述化合物的稳定性，膜渗透性和肿瘤特异性靶向。具有高PBD结合亲和力和特异性的化合物将转化为磷酸酶不敏感的、不可水解的p-Thr模拟物（Pmab）形式，然后进一步修饰以产生Ala-或Val-酯缀合的膦二酰胺前药。后一种修饰不仅是为了消除双阴离子膦酸部分的电负性以更好地跨细胞渗透，而且还将化合物靶向高度活性的PepT 1转运蛋白以有效地细胞摄取。对于在培养细胞水平上表现出抗Plk 1 PBD活性的化合物，我们将研究添加肿瘤靶向的RGD基序是否有助于化合物的肿瘤特异性递送。作为该项目的第三个目标，我们将确定所得化合物在小鼠肿瘤模型中抑制Plk 1依赖性细胞增殖活性的效力和选择性。由于胃肠道的化学和酶条件苛刻，小鼠尾静脉注射将是复方给药的选择。由于肽衍生的抑制剂通常表现出物理化学缺陷，我们还将采取补充方法，通过筛选NCI天然产物提取物储存库或通过进行计算机筛选来分离PBD抑制化合物或部分。分离的小分子化合物与上述PLHSpT衍生的抑制剂一起的结构分析和计算机建模将允许我们对后者抑制剂进行位点特异性替换和修饰，以实现增强的体内稳定性和生物利用度。为此，我们将带来我们的合作者的专业知识，从体外高通量筛选和基于细胞的测定（James McMahon博士），计算机筛选和计算机建模（Marc Nicklaus博士），以及X射线晶体学（Alex Wlodawer和Michael Yaffe博士）。这种多方面方法的最终目标是产生一类新的单特异性抗Plk 1治疗剂，这些治疗剂具有治疗人类各种癌症的潜力。