Inhibitors of B-raf through the Dimerization Interface

 描述（由申请人提供）：尽管B-Raf抑制剂（如维罗非尼）在临床上取得了成功，但在接受治疗的患者中出现了耐药性，从而抑制了对该方法的最初热情。对耐药机制的进一步研究提供了相当多的见解，并确定了一个主要原因是“矛盾的MEK/ERK信号传导”。研究表明，虽然维罗非尼有效抑制B-Raf V600 E，但在WT B-Raf同源二聚体和异源二聚体以及激活Ras突变的情况下，它导致二聚体中其他配偶体的激酶激活，从而刺激下游途径而不是抑制下游途径。因此，需要进一步努力以其他阻止下游信号传导的方式补充突变V600 E激酶的抑制。Brummer小组最近研究了野生型、致癌和药物抑制的B-Raf的二聚化的重要性，并表明二聚体界面（DIF）中的保守残基对表达致癌Ras的细胞中野生型B-Raf的信号传导潜力起重要作用。此外，B-Raf同源二聚体和异源二聚体的形成是基于不同的结构要求，并且对于Ras/Raf-1介导的药物结合B-Raf的反常MEK/ERK活化是必不可少的。这些发现不仅确定DIF作为潜在的治疗靶点（在Ras驱动的肿瘤中）或避免矛盾的MEK/ERK激活，而且还为DIF介导的反式激活的Raf激活的变构模型提供了强有力的支持。McInnes实验室设计了Raf二聚化的肽抑制剂，最近与Brummer小组合作，以证明它们作为致癌Ras信号传导事件的化学生物学探针的潜力。在实现阻断MEK/ERK信号传导的矛盾激活的概念证明之后，这样的抑制剂可以被认为是开发更多药物样化合物作为癌症治疗剂的先导。我们建议合成一个肽抑制剂库，以探索截短、取代和构象稳定方法对B-Raf DIF肽序列抑制二聚化事件的影响。肽工具化合物的开发和随后的细胞实验将使我们能够实现进一步理解Raf二聚化事件如何有助于Ras/Raf/MEK/ERK通路中信号的传播的目标，同时为在通过MEK/ERK的矛盾激活对维罗非尼耐药的肿瘤的背景下利用这些提供基础。

 DESCRIPTION (provided by applicant): Despite the clinical success of B-Raf inhibitors like vemurafenib, resistance has emerged in treated patients thereby dampening the initial enthusiasm for this approach. Further investigation into resistance mechanisms has provided considerable insights and determined that a major cause results from "paradoxical MEK/ERK signaling". Studies have demonstrated that while vemurafenib inhibits B-Raf V600E potently, in the context of WT B-Raf homodimers and heterodimers and activating Ras mutations, it leads to kinase activation of the other partner in the dimer thereby stimulating the downstream pathway rather than inhibiting it. Further efforts are therefore required to complement inhibition of the mutant V600E kinase with other ways of preventing downstream signalling. The Brummer group recently investigated the importance of dimerization for wildtype, oncogenic and drug inhibited B-Raf and showed that conserved residues in the dimer interface (DIF) play an important role for the signaling potential of wildtype B-Raf in cells expressing oncogenic Ras. Furthermore, the formation of B-Raf homo- and heterodimers is based on distinct structural requirements and is essential for Ras/Raf-1 mediated paradoxical MEK/ERK activation by drug-bound B-Raf. These findings identify not only the DIF as potential therapeutic target (in Ras-driven tumors) or to avoid paradoxical MEK/ERK activation, but also lend strong support to an allosteric model of Raf activation by DIF mediated transactivation. The McInnes laboratory has designed peptidic inhibitors of Raf dimerization and has recently initiated a collaboration with the Brummer group in order to demonstrate their potential as chemical biology probes of oncogenic Ras signaling events. After achieving proof of concept of blocking paradoxical activation of MEK/ERK signalling, such inhibitors could then be considered as leads for the development of more drug-like compounds as cancer therapeutics. We propose to synthesize a library of peptidic inhibitors to explore the effects of truncation, substitution and conformational stabilization approaches on the inhibition of dimerization events by the B-Raf DIF peptide sequence. The development of peptidic tools compounds and subsequent cellular experiments will enable to us to achieve the objectives of further understanding how Raf dimerization events contribute to propagation of signals in the Ras/Raf/MEK/ERK pathway while providing the basis for exploiting these in the context of tumors resistant to vemurafenib through paradoxical activation of MEK/ERK.