Development of Peptidomimetic Regulators of Myc function

Myc蛋白是通过激活和抑制转录来显著改变基因表达的转录因子。在人类中有三种Myc蛋白家族成员（c-Myc、N-Myc、L-Myc），它们都在癌症中异常表达。抑制Myc是一种有效的治疗策略，但开发直接靶向Myc蛋白的临床化合物的努力已经失败。c-Myc、N-Myc和L-Myc具有介导与关键伴侣蛋白相互作用的序列同源性区域。Myc反式激活结构域（Myc transactivation domain，MYC）本质上是无序的，如圆二色性和NMR光谱所报道的，但是存在瞬时二级结构元件，在某些情况下，其在与结合伴侣的复合物中变得稳定。Myc蛋白的几种相互作用伴侣在转录调控中相互作用并介导Myc的转录激活（或抑制）功能，如WDR 5和CDK 9。其他结合伴侣调节Myc的泛素化和降解，如FbxW 7和Aurora-A。N-Myc和Aurora-A之间的复合物的结构揭示了N-Myc蛋白的一部分折叠成螺旋，其包装在激酶的表面上。Aurora-A激酶调节有丝分裂进入和有丝分裂纺锤体组装，是一个有前途的抗癌治疗靶点。微管相关蛋白TPX 2通过结合两个位点激活Aurora-A，如Aurora-A-TPX 2复合物的X射线晶体结构所示。在这个博士生项目中，我们将开发基于肽模拟的关键Myc相互作用抑制剂。我们还将开发TPX 2肽模拟物，并研究它们对Aurora-A的结合亲和力。我们将利用最先进的方法来限制肽的生物活性构象;简单地说，通过可逆反应的肽轴承明智地放置硫醇侧链（Cys/ hCys）与二溴马来酰亚胺。约束肽被认为在蛋白水解/血清稳定性、由于预组织和细胞摄取性质而增强的靶亲和力方面是有利的。此外，我们还将利用这种能力的背景下的PTM（如磷酸化）的Myc肽，以评估有序/无序转换的蛋白质-蛋白质相互作用网络的作用。这将使相互作用的功能研究，例如，以确定在细胞周期中的特定点的相互作用的破坏的后果。开发的试剂将用于靶向验证实验，以探测Myc的哪些相互作用对癌细胞的存活至关重要，但在正常细胞类型中是不可或缺的。除了基本的洞察力，这个学生将提供对Myc的调节，确定一个潜在的战略，针对Myc，代表了第一阶段，在扩大该项目到药物发现。

Myc proteins are transcription factors that markedly alter gene expression through both activation and repression of transcription. There are three Myc protein family members in humans (c-Myc, N-Myc, L-Myc), which are all aberrantly expressed in cancers. Inhibition of Myc is a validated therapeutic strategy, but efforts to develop clinical compounds that target Myc proteins directly have failed. c-Myc, N-Myc and L-Myc have regions of sequence homology that mediate interactions with critical partner proteins. The Myc transactivation domain (TAD), is intrinsically disordered, as reported by circular dichroism and NMR spectroscopy, but there are transient secondary structure elements that, in some cases, become stable in complex with binding partners. Several interacting partners of Myc proteins interact in the TAD and mediate transcriptional activation (or repression) functions of Myc, such as WDR5 and CDK9. Other binding partners regulate the ubiquitination and degradation of Myc, such as FbxW7 and Aurora-A. The structure of the complex between N-Myc and Aurora-A, revealed that part of the N-Myc protein folds into a helix that packs against the surface of the kinase. Aurora-A kinase regulates mitotic entry and mitotic spindle assembly and is a promising target for anticancer therapy. The microtubule-associated protein TPX2 activates Aurora-A through binding to two sites, as shown by the X-ray crystal structure of the Aurora-A-TPX2 complex. In this PhD studentship we will develop peptidomimetic-based inhibitors of key Myc interactions. We will also develop TPX2 peptidomimetics and investigate their binding affinity for Aurora-A. We will exploit state of the art methodology developed to constrain peptides in a bioactive conformation; briefly through reversible reaction of peptides bearing judiciously placed thiol side-chains (Cys/ hCys) with dibromomaleimide. Constrained peptides are considered advantageous in terms of proteolytic/serum stability, enhanced target affinity due to preorganisation and cell-uptake properties. Moreover, we will also exploit this capability in the context of PTMs (e.g. phosphorylation) within the Myc peptides to evaluate the role of order/disorder transitions on the protein-protein interaction network. This will enable functional studies on the interactions, for example to determine the consequences of disruption of the interaction at a specific point in the cell-cycle. The developed reagents will be used in target validation experiments to probe which of the interactions of Myc are critical for the survival of cancer cells, but dispensible in normal cell types. Beyond the fundamental insight this studentship will deliver on regulation of Myc, Identification of a potential strategy for targeting Myc, represents the first stage in expanding the project into drug discovery.