PLEKHA7 A Novel Target for Mutant KRAS Therapy

PLEKHA7 突变 KRAS 治疗的新靶点

基本信息

批准号：
9301505
负责人：
GARTH POWIS
金额：
$ 62.41万
依托单位：
SANFORD BURNHAM PREBYS MEDICAL DISCOVERY INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-08-06 至 2020-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9301505
关键词：
Adherens Junction Anchorage-Independent Growth Automobile Driving Binding Binding Proteins CNKSR1 gene Cancer Cell Growth Cancer Patient Cell Line Cell membrane Cell physiology Cells Characteristics Colon Carcinoma Crystallization Cytoskeleton Disease E-Cadherin Epithelial Cells Event Family GTP Binding Genes Growth Guanosine Triphosphate Human Immunoprecipitation KRAS2 Gene Mutation KRAS2 gene Lead Legal patent Ligands Link Lipids Malignant Neoplasms Medical Membrane Membrane Proteins Microtubules Molecular Molecular Target Monomeric GTP-Binding Proteins Mutate Mutation Neoplasm Metastasis New Agents Normal Cell Oncogenes Oncogenic PH Domain Parents Patients Pharmaceutical Preparations Pharmacology Phenotype Phosphatidylinositols Plasma Proteins Play Point Mutation Positioning Attribute Property Proteins RAS inhibition Regulation Roentgen Rays Role Signal Pathway Signal Transduction Signaling Protein Site Small Interfering RNA Structure Surface Techniques Tight Junctions Validation Work Zinc Fingers base cancer cell cancer type carcinogenesis cell motility colon cancer cell line design effective therapy genetic regulatory protein imaging study in vivo inhibitor/antagonist knock-down member mutant novel novel strategies novel therapeutics preclinical study protein folding protein function public health relevance scaffold small molecule small molecule inhibitor therapy resistant tumor tumor growth

项目摘要

DESCRIPTION (provided by applicant): The KRAS oncogene is found in 25% of patient tumors across many cancer types. Despite extensive effort since its discovery over 40 years ago there is no effective treatment for mutated KRAS, and an estimated 350,000 patients in the US in 2014 with mutated KRas in their tumors will die of their disease. Mutated KRAS plays a critical role in driving tumor growth and resistance to therapy. Its effects are so powerful that i overrides the activity of many of the new molecularly targeted signaling drugs being developed for cancer today. Thus, finding new agents that inhibit the effects of mutated KRas is a critical unmet need in cancer today. However, intracellular signaling by wild type KRas protein controls many aspects of normal cell function, so that a therapy directed at inhibiting mutant KRas should ideally leave wild type KRas function unaffected. Using a global siRNA functional screen and isogenic cell line pairs of mutant or wild type KRas we identified PLEKHA7 (pleckstrin homology domain containing, family A7) as a protein that when knocked down inhibits the proliferation of colon cancer cells with mutant KRas, but remarkably not wild type KRas cells. Other aspects of the mutant KRas phenotype were also inhibited including anchorage independent (3D) growth, cell invasion, and in vivo tumor growth. We have also shown that PLEKHA7 knockdown decreases the active (GTP bound) form of mutant but not of wild type KRas, with inhibition of downstream mutant KRas signaling. PLEKHA7 is normally found in the adherens junction of normal epithelial cells. In cancer cells it is found in plasma membrane tight junctions but its function is not known. PLEKHA7 is a member of a group of signaling proteins containing a distinctive 3D protein fold, the pleckstrin homology (PH)-domain, that binds to membrane phosphoinositides to position the parent proteins at specific sites on the membrane important for their function. Our previous studies have shown that PH domains can be selectively drugged by small molecules, thus inhibiting the signaling function of the proteins. We have evidence that in cancer cells PLEKHA7 is associated with proteins of the plasma membrane-bound KRas signaling nanocluster. We thus hypothesize that PLEKHA7 selectively regulates mutated KRas activity in the signaling nanocluster, and therefore that the PH- domain of PLEKHA7 is a target for small molecules inhibitors as pharmacological probes for mechanistic studies of PLEKHA7 function, as well as selectively blocking the growth of mutated KRas cancer cells as potential therapy. This represents a new paradigm for attacking Ras via inhibition of associated regulatory signaling nanocluster proteins. The objectives of our study are: 1) to investigate the mechanism for PLEKHA7's ability to selectively inhibit mutated KRas; 2) to conduct structural studies of the PLEKHA7 PH domain and two potential "hinge" regions, to delineate PLEKHA7's function in associating with the plasma membrane and other proteins of the KRas signaling nanocluster; and 3) to identify small molecule inhibitors of the PLEKHA7 PH- domain as pharmacological probes to study the role of PLEKHA7 in KRas regulation, and as leads for potential agents to treat mutated KRas tumors.

描述（由适用提供）：在许多癌症类型的患者肿瘤中发现KRAS癌基因。尽管自从40年前发现以来，却没有有效的KRAS治疗方法，而2014年美国估计有35万名患者患有肿瘤中的KRAS突变的KRAS将死于其疾病。突变的KRAS在推动肿瘤生长和对治疗的抵抗力中起着至关重要的作用。它的作用是如此强大，以至于我覆盖了当今为癌症开发的许多新分子信号药物的活性。这是找到抑制突变KRAS影响的新药物是当今癌症的至关重要的需求。但是，野生型KRAS蛋白的细胞内信号传导控制正常细胞功能的许多方面，因此一种针对抑制突变体Kras的治疗应理想地使野生型KRAS功能不受影响。使用突变体或野生型KRAS的全局siRNA功能筛选和等源细胞系对，我们确定了plekha7（含有含有家族A7的pleckstrin同源性结构域）为蛋白质，当敲低时，抑制了与突变体KRA的结肠癌细胞的增殖时，但不是野生型Kras细胞。突变体KRAS表型的其他方面也受到抑制，包括锚定独立（3D）生长，细胞侵袭和体内肿瘤生长。我们还表明，PLEKHA7敲低降低了突变体的活性（GTP结合）形式，而不是野生型KRAS，并抑制了下游突变体KRAS信号传导。 Plekha7通常在正常上皮细胞的粘附连接中发现。在癌细胞中，它在质膜紧密连接处发现，但其功能尚不清楚。 plekha7是一组含有独特的3D蛋白折叠的信号蛋白的成员，该蛋白折叠（PLECKSTRIN同源性（pH）域）与膜磷酸肌醇结合，以将父蛋白定位在对其功能的特定位点上的特定位点。我们以前的研究表明，可以通过小分子选择pH结构域，从而抑制蛋白质的信号传导功能。我们有证据表明，在癌细胞中，plekha7与质膜结合的KRAS信号纳米簇的蛋白质有关。因此，我们假设plekha7选择性调节信号纳米群中突变的KRAS活性，因此，plekha7的ph域是小分子抑制剂作为PLEKHA7功能的机械研究的小分子抑制剂的目标，并选择性地阻断了突变的Kras癌细胞的生长，并选择了潜在的癌细胞的生长。这代表了通过抑制相关的调节信号纳米簇蛋白来攻击RA的新范式。我们研究的目标是：1）研究Plekha7有选择性抑制突变KRAS的能力的机制； 2）进行Plekha7 pH结构域和两个潜在的“铰链”区域进行结构研究，以描绘plekha7的功能与质膜和KRAS信号纳米簇的其他蛋白质相关联； 3）确定Plekha7 ph域的小分子抑制剂是研究Plekha7在KRAS调节中的作用的药物问题，并作为治疗突变的KRAS肿瘤的潜在药物的铅。