Aryl Hydrocarbon Receptor Modulators for the Treatment of Hepatocellular Carcinom

用于治疗肝细胞癌的芳基烃受体调节剂

• 批准号: 8050195
• 负责人: Siva Kumar Kolluri
• 金额: $ 18.28万
• 依托单位: OREGON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2013-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8050195
• 关键词: AHR gene; Androgen Antagonists; Antineoplastic Agents; Apoptosis; Aryl Hydrocarbon Receptor; Binding; Biological; Biological Assay; Cell Culture Techniques; Cell Cycle Regulation; Cell Death; Cell Nucleus; Cell Proliferation; Cells; Computer Simulation; Coupled; Cytosol; DNA Binding; Databases; Development; Dioxins; Evaluation; Family member; Fingerprint; Flutamide; Future; Gene Expression; Gene Expression Profiling; Gene Targeting; Genetic Transcription; Growth; Health; Homology Modeling; Human; Inhibition of Apoptosis; Inhibition of Cancer Cell Growth; Lead; Ligand Binding; Ligands; Liver; Malignant Neoplasms; Malignant neoplasm of liver; Malignant neoplasm of prostate; Mediating; Mediator of activation protein; Modeling; Molecular Profiling; Molecular Target; Mus; Prevention; Primary carcinoma of the liver cells; Protein Family; Proteins; Receptor Activation; Receptor Signaling; Reporter; Research; Role; Screening procedure; Structural Models; Structure; Testing; Tetrachlorodibenzodioxin; Therapeutic; Therapeutic Intervention; Toxic effect; Transcriptional Activation; Tumor Suppression; Tumor Suppressor Proteins; Virtual Library; activating transcription factor; aryl hydrocarbon receptor ligand; base; cancer cell; cancer therapy; hepatoma cell; human AHR protein; in vivo; interest; malignant breast neoplasm; novel; programs; response; single-minded protein; three dimensional structure; tumor growth; tumor xenograft; virtual

DESCRIPTION (provided by applicant): The Aryl Hydrocarbon Receptor (AhR) is a ligand-activated transcription factor. Upon binding to a ligand, the AhR translocates from the cytosol to the nucleus, where it mediates the transcription of a wide range of gene targets. Aside from the AhR's role as the mediator of toxicity of various dioxins, recent studies have identified roles for the AhR in apoptosis, cell cycle regulation, and tumor suppression, all of which make the AhR a potentially attractive molecular target for development of novel cancer therapeutics. We hypothesize that certain selective AhR modulators (SAhRMs) are capable of initiating biological responses that should prove useful for prevention and/or treatment of hepatocellular carcinoma. We propose to identify SAhRMs by in silico structure-based Virtual Ligand Screening (VLS) coupled with cell- based AhR transcriptional activation assays. SAhRMs will be screened further for their ability to induce AhR- dependent cell death and/or inhibition of cancer cell proliferation. The availability of numerous well- characterized AhR expressing and non-expressing cell culture models in our lab will enable us to evaluate the functional consequences of AhR activation and the anticancer effects of sAhRMs in hepatoma cells. The Specific Aims proposed in this application are described below. 1. Identify novel AhR modulators that have the potential for treatment of cancer We will use in silico structure-based VLS for identifying novel alternative ligands of the AhR. VLS utilizes a three dimensional structure of a protein or homology model to identify potential binders of a target protein. The Per-Arnt-Sim (PAS)-B domain of AhR is essential for ligand binding. As the AhR ligand binding pocket is not yet crystallized, we have generated and refined mouse and human AhR structural models based on the AhR's homology with existing PAS protein family members whose three-dimensional structures have been solved. These homology models of the AhR will be used to screen both synthetic and natural compound databases by VLS to identify potential binders of the AhR. The effects of the newly identified compounds on AhR activation will be tested by AhR-dependent reporter assays and evaluation of AhR- dependent induction of endogenous target genes. Preliminary screening of a compound library by VLS using the AhR PAS domain homology model described above has already resulted in the identification of some compounds of interest including the clinically used antiandrogen flutamide that promoted transcriptional activation of AhR, induction of AhR- regulated target genes and AhR-dependent inhibition of cancer cell growth. These results strongly support of the feasibility of this exploratory research. 2. Test the compounds identified by VLS for AhR selectivity and potential anticancer effects The compounds identified by VLS will be tested for their ability to directly bind to the AhR. Molecules that are able to bind to the AhR will be tested for their effects on AhR DNA binding and inducing AhR-specific transcriptional activity in liver cancer cells. AhR ligands that induce cell death and/or antiproliferative effects in an AhR-dependent manner in liver cancer cells will be prioritized for further evaluation. 3. Shortlist compounds for future in vivo testing by gene expression profiling We will compare the gene expression induced by SAhRMs to distinguish them from the transcriptional program induced by the classical AhR ligands such as TCDD using human and mouse microarrays. Compounds with expression profiles that differ significantly from that of TCDD, such as our initial lead compounds (flutamide and C-24), and that induce growth inhibition/apoptosis via AhR will be given priority for future in vivo studies. SAhRMs with gene expression profiles highly similar to that of TCDD will not be pursued as anti-cancer agents. The proposed studies are timely and important in the context of developing therapeutic intervention for the prevention and/or treatment of hepatoma via an intriguing molecular target, AhR. Excitingly, the SAhRMs that will be identified by these studies may also be effective against other cancers. We will evaluate some of short-listed SAhRMs for AhR-dependent inhibition of tumor growth in mouse xenografts tumor studies in the future.

描述(申请人提供)：芳烃受体(AhR)是一种配体激活的转录因子。当与配体结合时，AhR从胞浆转移到细胞核，在那里它介导了广泛的基因靶标的转录。除了AhR作为多种二恶英毒性的介体外，最近的研究已经确定了AhR在细胞凋亡、细胞周期调节和肿瘤抑制中的作用，所有这些都使AhR成为开发新的癌症治疗药物的潜在分子靶点。我们假设某些选择性AhR调节剂(SAhRms)能够启动生物学反应，这应该被证明对预防和/或治疗肝细胞癌是有用的。我们建议通过基于电子结构的虚拟配体筛选(VLS)和基于细胞的AhR转录激活分析来识别SAhRms。SAhRms将进一步筛选其诱导AhR依赖的细胞死亡和/或抑制癌细胞增殖的能力。我们实验室建立了大量表达AhR和不表达AhR的细胞培养模型，这将使我们能够评估AhR激活的功能后果和sAhRms在肝癌细胞中的抗癌作用。本申请中提出的具体目标如下所述。1.鉴定具有治疗癌症潜力的新型AhR调节剂我们将在基于硅结构的VLS中用于鉴定AhR的新型替代配体。VLS利用蛋白质的三维结构或同源模型来识别目标蛋白质的潜在结合蛋白。AhR的Per-Arnt-Sim(PAS)-B结构域是配体结合所必需的。由于AhR配体结合口袋尚未结晶，我们根据AhR与现有PAS蛋白家族成员的同源性建立和完善了小鼠和人AhR结构模型，其三维结构已经得到解决。这些AhR的同源模型将被VLS用于筛选合成化合物数据库和天然化合物数据库，以确定AhR的潜在结合子。新鉴定的化合物对AhR激活的影响将通过AhR依赖的报告分析和评估AhR依赖的内源性靶基因的诱导来测试。VLS使用上述AhR PAS结构域同源模型对化合物文库的初步筛选已经导致了一些感兴趣的化合物的鉴定，包括临床使用的抗雄激素氟他胺，它促进AhR的转录激活，诱导AhR调节的靶基因，并依赖于AhR抑制癌细胞生长。这些结果有力地支持了本探索性研究的可行性。2.测试VLS鉴定的化合物的AhR选择性和潜在的抗癌作用将测试VLS鉴定的化合物直接与AhR结合的能力。能够与AhR结合的分子将被测试它们对AhR DNA结合的影响，并在肝癌细胞中诱导AhR特异性转录活动。以AhR依赖的方式在肝癌细胞中诱导细胞死亡和/或抗增殖作用的AHR配体将被优先考虑进行进一步评估。3.利用基因表达谱进行体内实验的候选化合物我们将利用人和小鼠微阵列比较SAhRms诱导的基因表达，以区分它们与经典的AhR配体如TCDD诱导的转录程序。表达谱与TCDD显著不同的化合物，如我们最初的先导化合物(氟他胺和C-24)，以及通过AhR诱导生长抑制/凋亡的化合物将被优先用于未来的体内研究。基因表达谱与TCDD高度相似的SAhRM将不会被用作抗癌药物。在开发通过耐人寻味的分子靶点AhR预防和/或治疗肝癌的治疗干预措施的背景下，拟议的研究是及时和重要的。令人兴奋的是，这些研究将确定的SAhRM也可能对其他癌症有效。我们将在未来的小鼠移植瘤研究中评估一些入围的SAhRms对AhR依赖的肿瘤生长的抑制作用。