New Ways of Targeting K-Ras

靶向 K-Ras 的新方法

基本信息

批准号：
8956333
负责人：
FRANK PATRICK MCCORMICK
金额：
$ 93.29万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-04-01 至 2023-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8956333
关键词：
Adverse effects Affect Antibodies Area Binding Binding Proteins Biochemical Calmodulin Cell Count Cells Complex Development Disease Drug resistance Genetic Genetic Transcription Goals Human Individual Intention KRAS2 gene LIF gene MAP Kinase Gene Malignant Neoplasms Malignant neoplasm of pancreas Modeling Monoclonal Antibodies NF1 gene Natural Products Neoplasm Metastasis Neurofibromatoses Neurofibromatosis Type 1 Protein Oncogenic Oral Administration Pathway interactions Phenotype Phosphorylation Play Preclinical Testing Property Proteins Regulation Research Role Safety Serine Signal Transduction Stem cells Testing Tumor Suppressor Proteins analog base beta catenin bone cancer cell gemcitabine in vivo mouse model mutant neoplastic cell novel pancreatic neoplasm pre-clinical prevent prostratin public health relevance ras Proteins research clinical testing research study tumor tumor initiation

项目摘要

DESCRIPTION (provided by applicant): We propose three related area of research, each focused on a major aspect of Ras signaling. The first attempts to solve an important question relating to the NF1 gene, now recognized as a major tumor suppressor in human cancer as well as genetic basis for neurofibromatosis, a disease that affects 1 in 3500 people worldwide. Using biochemical approaches as well as cell-based screens, we will try and identify proteins or other cellular factors that regulate neurofibromin's ability to regulate Ras. Recently, we identified the first bone fide neurofibromin binding proteins other than Ras, the Spred proteins that are essential for neurofibromin's activity. We will build on this discovery to determine how the neurofibromin/Spred complex is assembled and regulated, and, in addition, we will perform unbiased screens for factors that regulate Ras through neurofibromin, or regulate Ras-independent functions of this highly conserved protein. Second, we will analyze individual mutants of K-Ras for their requirement for upstream signaling, and for the specific activation of downstream pathways. We will do this, as in the previous section, by combining analysis of biochemical properties of the proteins themselves, and analysis of downstream pathways. We will focus on known pathways that are clearly activated differently by different oncogenic forms of Ras, such as the Raf MAPK pathway, and unbiased analysis of signaling differences that are revealed by analysis of individual proteins in a clean, isogenic background. Finally, we have discovered that K-Ras promotes a stem-cell like phenotype in cancer cells that enables them to form tumors very efficiently from small numbers of cells, to metastasize and to become drug resistant. These phenotypes are caused by K-Ras binding to calmodulin and suppressing non-canonical wnt signaling, while increasing transcription from beta-catenin and other factors involved in stem-cell signaling. One of these factors that is produced by K-Ras cancers and plays a major role in the stem-cell phenotype is Leukemia Inhibitory Factor (LIF). We have found that neutralizing LIF prevents tumor initiation in vivo and sensitizes pancreatic tumor cells to gemcitabine, so that established tumors can be completely eradicated. We propose to undertake pre-clinical analysis of humanized anti-LIF monoclonal antibodies, using several models of pancreatic cancer, with the intention of moving these antibodies into clinical testing based on the results of these experiments. Binding of K-Ras to calmodulin is prevented by phosphorylation on serine-181, by PKC. The non-tumor promoting natural product, prostratin, activates PKC in vivo and disrupts K-Ras-calmodulin interaction. Oral administration of prostratin prevents development of pancreatic cancers in mouse models, with no obvious side effects. Several analogs of prostratin have been synthesized: we will test all of these, and make more. We will identify which form of PKC is involved in K-Ras phosphorylation and perform thorough preclinical testing for safety and efficacy of the most promising analogs, so that, if successful, we will advance one of these towards clinical testing.

描述（由适用提供）：我们提出了三个相关的研究领域，每个领域都集中在RAS信号的主要方面。第一次尝试解决了与NF1基因有关的重要问题，该问题现在被公认为是人类癌症中的主要肿瘤抑制剂以及神经纤维瘤病的遗传基础，神经纤维瘤病是一种影响全球3500人中有1人的疾病。使用生化方法以及基于细胞的筛选，我们将尝试鉴定蛋白质或其他调节神经纤维蛋白调节RAS能力的细胞因子。最近，我们确定了除RAS以外的其他骨FIDE神经纤维蛋白结合蛋白，SPRED蛋白对于神经纤维蛋白的活性至关重要。我们将基于这一发现，以确定如何组装和调节神经纤维蛋白/SPRED复合物，此外，我们将对通过神经纤维蛋白调节RAS的因素执行无偏见，或调节该高度组成的蛋白质的RAS无关功能。其次，我们将分析K-RAS的个别突变体，我们将专注于已知的途径，这些途径通过不同的RAS的致癌形式（例如RAF MAPK途径）明显不同地激活，以及对信号差异的无偏分析，这些分析通过清洁，异源背景中单个蛋白质的分析所揭示的信号差异。最后，我们发现K-RAS促进了癌细胞中的干细胞类似的表型，从而使它们能够从少量细胞，转移并变得耐药性地形成肿瘤。这些表型是由K-RAS与钙调蛋白结合并抑制非典型WNT信号传导引起的，同时增加了β-蛋白的转录以及与干细胞信号传导有关的其他因素。这些因素是由K-RAS癌所产生的因素之一，在干细胞表型中起主要作用是白血病抑制因子（LIF）。我们发现中和LIF可以防止体内肿瘤的启动，并感觉到胰腺肿瘤细胞到吉西他滨，可以完全放射良好的肿瘤。我们建议使用几种胰腺癌模型对人性化抗LIF单克隆抗体进行临床前分析，以根据这些实验的结果将这些抗体转移到临床测试中。 PKC在丝氨酸-181上的磷酸化阻止K-RAS与钙调蛋白的结合。促进天然产物Prestratin的非肿瘤在体内激活PKC并破坏K-Ras-Calmodulin的相互作用。口服prostratin可以防止小鼠模型中胰腺癌的发展，没有明显的副作用。 prstratin的几种类似物已经合成：我们将测试所有这些，并进行更多。我们将确定哪种形式的PKC参与K-RAS磷酸化，并对最有希望的类似物的安全性和有效性进行彻底的临床前测试，因此，如果成功，我们将将其中之一推向临床测试。