Novel Chemical Biology Probes based on Selective Inhibitors of the Polo-Box Domain of PLK1

Polo-box结构域（PBD）对PLK 1的有丝分裂功能至关重要，并且已显示为底物 募集和亚细胞定位。McInnes实验室已经取得了重大进展， 开发基于PBD抑制的PLK 1选择性抑制剂，其对PLK 3肿瘤具有最小活性 抑制子，并保留了针对PLK 1位点突变体活性，所述突变体对临床使用的ATP具有抗性 竞争性抑制剂这些化合物将是进一步建立生物化学的高价值化学生物学探针 通过PBD以非ATP竞争性方式抑制PLK 1的表型后果， PBD作为抗有丝分裂药物靶点以及它们作为下一代治疗剂的潜力。这项建议是 在以下方面具有重要意义和创新性： 1)已经开发了一种称为REPLACE的创新策略来产生细胞周期特异性CDK 抑制剂作为抗肿瘤治疗剂。此外，McInnes实验室验证了这种策略， PLK 1 PBD揭示了通过REPLACE发现的片段连接的肽抑制剂， 相当可观的抗增殖活性和与靶抑制一致的表型。 2)靶向PBD的化学生物学探针对PLK 1具有高度选择性， 对PLK 3肿瘤抑制因子的最小亲和力。这对于进一步建立表型 选择性阻断PLK 1 PBD的后果，并且还避免PLK 3 在抗肿瘤治疗剂的开发中的抑制作用。我们已经证明， 有效的PLK 1 PBD结构域抑制剂对PLK 3具有最小的活性。 3)PBD靶向化合物将保留针对已经获得的肿瘤细胞的抗肿瘤活性。 对基于ATP的抑制剂的抗性。如上所述，单点突变体（C67 V）能够 使PLK 1对结构上不同的基于ATP的抑制剂具有抗性。针对非催化功能 PLK 1的抑制作用应导致较小的耐药选择压力，PBD抑制剂可用于 与基于ATP的抑制剂组合作为临床中PLK 1靶向的协同手段。 4)目前的先导化合物在处理的细胞中诱导重现PLK 1的表型 击倒。这与弱结合的小分子PBD抑制剂形成对比，据报道， 诱导部分效应，并且可能不具有期望水平的抗肿瘤活性。我们观察到的一些 表型是新的，提示后期突变，已知促进肿瘤选择性。 5)新的PBD抑制剂将是肿瘤选择性化合物。最近发现PLK 1抑制是 在PTEN缺陷型、突变型Ras和p53缺陷型癌症中， 肿瘤选择性化合物非肽抑制剂将用于细胞研究，以确认肿瘤细胞 特异性、PLK 1选择性和细胞表型与靶向抑制一致。

The Polo-box domain (PBD) is critical for mitotic functions of PLK1 and has shown to be required for substrate recruitment and sub-cellular localization. The McInnes laboratory has made significant progress toward the development of PLK1 selective inhibitors based on PBD inhibition that have minimal activity on the PLK3 tumor suppressor and which retain activity against PLK1 site mutants that are resistant to clinically utilized ATP competitive inhibitors. These compounds will be high value chemical biology probes for further establishing the phenotypic consequences of inhibiting PLK1 in a non-ATP competitive fashion through the PBD, in validating the PBD as an anti-mitotic drug target and in their potential as next generation therapeutics. This proposal is significant and innovative with respect to the following: 1) An innovative strategy called REPLACE has been developed to generate cell cycle specific CDK inhibitors as anti-tumor therapeutics. Furthermore the McInnes laboratory validated this strategy for the PLK1 PBD revealing that fragment ligated peptidic inhibitors discovered though REPLACE have respectable anti-proliferative activity and phenotypes consistent with target inhibition. 2) Chemical biology probes that target the PBD will be highly selective for PLK1 and possess minimal affinity for the PLK3 tumor suppressor. This is important to further establish the phenotypic consequences of blocking the PLK1 PBD selectively and also to avoid deleterious effects of PLK3 inhibition in the development of anti-tumor therapeutics. We have already demonstrated that our most potent PLK1 PBD domain inhibitors have minimal activity against PLK3. 3) PBD targeted compounds will retain antitumor activity against tumor cells that have acquired resistance to ATP-based inhibitors. As described above, a single point mutant (C67V) is capable of rendering PLK1 resistant to structurally diverse ATP-based inhibitors. Targeting non-catalytic functions of PLK1 should result in less selective pressure for resistance and PBD inhibitors might be used in combination with ATP-based inhibitors as a synergistic means of PLK1 targeting in the clinic. 4) Current lead compounds induce a phenotype in treated cells that recapitulates a PLK1 knockdown. This stands in contrast to weakly binding small molecule PBD inhibitors reported that only induced partial effects, and may not possess a desirable level of anti-tumor activity. Some of our observed phenotypes are novel and suggestive of anaphase catastrophe, known to promote tumor selectivity. 5) Novel PBD inhibitors will be tumor selective compounds. Recent discoveries that PLK1 inhibition is synthetically lethal in PTEN deficient, mutant Ras and p53 deficient cancers will be exploited to generate tumor selective compounds. Non-peptidic inhibitors will be used in cellular studies to confirm tumor cell specificity, PLK1 selectivity, and cellular phenotypes consistent with on target inhibition.