Structure-based In silico Screen for Small Molecule Inhibitors of Ets-1 Activity

描述(申请人提供)：炎症过程有助于包括类风湿性关节炎(RA)在内的自身免疫性疾病中导致组织破坏的病理事件。对于RA患者，能够阻断TNF1和IL-1的治疗药物的开发一直是重要的治疗里程碑，然而，相当多的患者未能对这些治疗产生反应，可能是因为他们天生无法抑制这种复杂疾病所需的其他途径。一些实验室建议，为了进一步了解类风湿关节炎的病理生理机制，需要识别、靶向和验证新的治疗靶点。一类潜在的靶点是细胞因子诱导的转录因子：核因子-k B、AP-1、C/EBP和Ets-1，它们都已在RA滑膜中检测到。最近的一些文献支持靶向转录因子(TF)的重要性，这将提供一种重新调节基因表达途径的机制，包括通过受控的变阻器“开关”而不是二元开关机制来调节细胞因子途径。此外，丰富的信息和对转录生物学和基因调控的更好理解支持了转录因子本身是治疗干预的潜在靶点。拟议的项目具有很高的创新性，将我们以翻译发现为基础的科学与这一新靶空间的体外和体内验证相结合，该空间是用于开发首例人型RA治疗的TF-DNA接口。这一协作和真正协同的应用涉及具有互补技能的研究人员，其优势在于通过基于核磁共振波谱的靶标验证和体外鉴定/评估来发现基于硅结构的小分子，并支持我们的方法来识别和开发特定靶向并抑制ETS-1与其序列特异性DNA启动子元件之间相互作用的新型小分子。这些研究人员拥有独特的优势和专业知识，当他们合作时，这为创造性的、“开箱即用”的思维和执行提供了一个重要的机会，支持这一申请的重要初步数据证明了这一点。具有明显活性的小分子，如那些已确定的小分子，代表着依赖于Tf的转录调控的诱人机会，为在Tf-DNA水平选择性抑制ETS Tf活性的治疗剂的开发提供了一种创新的策略。这一应用的重点是选择性地靶向Tf-DNA界面，这代表了一种依赖于Tf的转录调控ETS-1的新方法，并为选择性靶向该转录因子的治疗剂的开发提供了独特的机会。转录因子-DNA相互作用界面的小分子抑制为类风湿关节炎的转录治疗提供了一种有希望的范式转变，如核因子-kB(NF-kB)所尝试的途径特异性转录调控。通过丰富的信息和对转录生物学和基因调控的更好理解，包括核因子-kB、HIF112等在内的转录因子已成为治疗干预的新靶点。拟议的项目具有很高的创新性，将我们基于翻译发现的科学与新靶点空间的体外和体内验证相结合；TF-DNA接口用于识别和随后开发治疗RA的新疗法。转录因子是基因表达的既定调节因子，因此是各种生物过程所必需的，包括生长、分化和发育以及癌症和/或炎症等病理过程。序列特异性的Tf-DNA相互作用在空间和时间上都是受调控的，从而在蛋白质-DNA界面上产生了精致的特异性和选择性。选择性地靶向和抑制TF-DNA复合体的相互作用界面的能力代表了一种新的、高度特异的策略，用于重新编程在RA和癌症中解除调控的特定基因途径。与其他方法不同，包括聚酰胺、人工转录因子、锌指蛋白疗法，我们建议在电子计算机、虚拟筛选或高通量对接(HTD)中合作，以筛选大的公共可访问的化学储存库中的小分子，这些小分子选择性地靶向并从而抑制这种明确定义的分子界面。重要的是，在硅胶中，HTD非常适合于探索新的目标空间，如ETS-1 Tf-DNA相互作用界面。我们的中心假设是，靶向并破坏该界面的小分子将能够调节异常的基因转录，从而为类风湿关节炎提供一种新的治疗范例。这些在体内表现出显著活性的小分子支架将为类风湿关节炎的转录治疗提供平台。如果成功，这种方法代表着一个创新的和非常独特的机会，可以开发一种“人类第一”的治疗方法，通过靶向转铁蛋白的选择性/特异性界面来选择性地靶向和干扰下游转铁蛋白介导的基因表达。

DESCRIPTION (provided by applicant): Inflammatory processes contribute to the pathological events that lead to tissue destruction in autoimmune diseases including rheumatoid arthritis (RA). For patients suffering with RA the development of therapeutic agents that are capable of blocking TNF1 and IL-1 have been important therapeutic milestones, however a significant number of patients fail to respond to these therapies possibly due to their inherent inability to inhibit other pathways requisite in this complex disease. Several laboratories have suggested that in order to further understand the pathophysiologies of RA new therapeutic targets need to be identified, targeted and validated. One potential class of targets is the cytokine-induced transcription factors; NF-k B, AP-1, C/EBP, and ETS-1 all of which have been detected in RA synovium. Several recent publications support the importance of targeting transcription factors (TFs), which would provide a mechanism of re-regulating gene expression pathways including the cytokine pathway through a controlled, rheostat "switch" rather than a binary on/off mechanism. In addition, a wealth of information and a better understanding of transcriptional biology and gene regulation supports that TFs themselves are potential targets for therapeutic intervention. The proposed project is highly innovative, partnering our translational discovery-based science with in vitro and in vivo validation of this novel target space, the TF-DNA interface for use in the development of first in man type RA therapy. This collaborative and truly synergistic application involving investigators with complementary skills partners the strengths of in silico structure-based small molecule discovery with NMR spectroscopy based target validation and in vitro characterization/evaluation in support of our approach for identifying and developing novel small molecules that specifically target and inhibit the interaction interface between Ets-1 and its sequence specific DNA promoter element. These investigators have unique strengths and expertise, which when partnered provides a significant opportunity for creative, "out of the box" thought and execution as demonstrated by the significant preliminary data in support of this application. Small molecules with demonstrable activity such as those identified represent an attractive opportunity for TF-dependent transcriptional regulation, providing an innovative strategy for the "hit-through-lead" development of therapeutic agents that selectively inhibit ETS TF activity at the TF-DNA level. This application is focused on selectively targeting this TF-DNA interface, which represents a novel approach for TF-dependent transcriptional regulation of Ets-1 and provides a unique opportunity for the development of therapeutic agents selectively targeting this transcription factor. Small molecule inhibition of the TF-DNA interaction interface provides a promising paradigm shift in transcriptional therapy for RA through pathway specific transcriptional regulation as has been attempted for nuclear factor kappa B (NF-kB).Through a wealth of information and a better understanding of transcriptional biology and gene regulation, TFs including NF-kB, HIF112 and others have emerged as novel targets for therapeutic intervention. The proposed project is highly innovative, partnering our translational discovery-based science with in vitro and in vivo validation of novel target space; the TF-DNA interface for use in the identification and subsequent development of novel therapies for RA. TFs are established regulators of gene expression and as such are requisite for a variety of biological processes, including growth, differentiation and development as well as pathological processes such as cancer and/or inflammation. Sequence-specific TF-DNA interactions are spatially and temporally regulated, resulting in refined specificity and selectivity at the protein-DNA interface. The ability to selectively target and inhibit the interaction interface of the TF-DNA complex represents a novel, highly specific strategy for reprogramming specific gene pathways that are deregulated in RA and cancer. Unlike other approaches including; polyamides, artificial transcription factors, zinc finger protein therapeutics we have proposed to partner in silico, virtual screening or high throughput docking (HTD) to screen large publicly accessible chemical repositories for small molecules that selectively target and thus inhibit this well-defined molecular interface. Importantly, in silico HTD is ideally suited for the exploration of novel target space such as the Ets-1 TF-DNA interaction interface. Our central hypothesis is that small molecules that target and disrupt this interface would be capable of regulating aberrant gene transcription and would thus offer a novel paradigm of therapy for RA. Those small molecule scaffolds that demonstrate significant in vivo activity would provide a transcriptional therapy platform for RA. This approach, if successful, represents an innovative and very unique opportunity to develop a "first in man" therapeutic approach that will selectively target and disrupt downstream TF-mediated gene expression by targeting the selectivity/specificity interface of the TF.