Mechanisms of Kinase Function and Drug Resistance in Cancer

癌症中激酶功能和耐药性的机制

• 批准号: 8288696
• 负责人: Jianwei Che
• 金额: $ 34.29万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2015-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8288696
• 关键词: Acute Lymphocytic Leukemia; Adverse effects; Affect; Alleles; Architecture; Autoimmunity; Automobile Driving; B-Cell Development; B-Lymphocytes; Basic Amino Acids; Binding; Binding Proteins; Bone Marrow; CSNK1A1 gene; Cancer Etiology; Catalysis; Catalytic Domain; Cells; Chimeric Proteins; Chronic Myeloid Leukemia; Clinical; Clinical Trials; Computer Architectures; Cyclic GMP-Dependent Protein Kinases; Cytostatics; Data; Development; Disease; Drug Delivery Systems; Drug resistance; Electrostatics; Exclusion; Family; Follow-Up Studies; Funding; Generations; Gleevec; Hydrogen Bonding; Hyperactive behavior; Imatinib; In Vitro; Malignant Neoplasms; Mediating; Metabolism; Modeling; Molecular; Mus; Mutagenesis; Mutation; Oncogenes; Patients; Pharmaceutical Preparations; Phosphotransferases; Protein Kinase; Proteins; Publishing; Relapse; Resistance; Role; Serine; Side; Sodium Chloride; Splenomegaly; Structure; Test Result; Testing; Therapeutic; Therapeutic Uses; Threonine; Time; Triad Acrylic Resin; Tyrosine; Variant; base; casein kinase; cell growth; cell transformation; design; drug discovery; improved; in vitro activity; in vivo; in vivo Model; inhibitor/antagonist; innovation; insight; interdisciplinary approach; kinase inhibitor; lyn protein-tyrosine kinase; molecular dynamics; mutant; neoplastic cell; patient population; resistance mutation; small molecule; src-Family Kinases; success

DESCRIPTION (provided by applicant): Kinases are the second-largest drug-target family with 10 approved kinase inhibitor drugs and 50 compounds in clinical trials. Protein-kinase-domains are most frequently encoded by cancer-genes. Several cancer-driving mutations occur in their ATP-binding G-loops. The Abl-inhibitor Imatinib is a breakthrough-therapeutic for chronic-myelogenous-leukemia, but ~35% of the patients relapse due to accumulation of Imatinib-resistant Abl kinase-domain-mutations, particularly in the G-loop. Drug-resistance could thus become a major clincial problem as increasing patient populations are treated with kinase-inhibitor drugs. Using the Src-family protein tyrosine kinase Lyn as an experimentally very tractable example, we propose to implement and validate a multidisciplinary approach that first uses molecular dynamics (MD) simulations to relatively quickly identify mutations that affect catalysis and inhibitor interactions and can cause drug-resistance (Aim 1). Our approach next analyzes the activities, inhibitor-interactions and -resistance of the identified Lyn mutants in vitro and in vivo in Ba/F3 cells (Aim 2) or in Lyn-/- bone-marrow (Aim 3) to identify those mutations that are most relevant physiologically. Exclusion of uninformative mutants at each step minimizes experimental effort and maximizes relevance and likelihood of success. We consider this integrated approach to discover drug-resistance causing kinase mutations highly innovative, because it provides important insight that is usually only gained over much longer time periods and through the efforts of several labs. These studies follow up on our recently published finding that 58 eukaryotic kinases contain a conserved electrostatic salt-bridge across their G-loops that is essential for G-loop-stabilization, catalysis and ATP- or ATP-competitive inhibitor-binding. Salt-bridge- disruption in Bcr-Abl causes Imatinib-resistance. Our preliminary data suggest that in 31 kinases, including the Src, Abl, CK1 and CK2-families which all have important roles in cancer, the acidic salt-bridge-anchor also interacts electrostatically with a conserved polar-aromatic or basic amino-acid-side-chain embedded in a hydrophobic core. To test the hypothesis that this "triad interaction-network" architecture is essential for G-loop function and inhibitor-interactions, and that its disruption can cause drug resistance, we will analyze the effects of mutationally modulating the different components of the variant G-loop-triad-configurations in the exemplary kinases Lyn (Aims 1-3), Abl, CK1(2 and CK2a1 (Aim 4). To keep Aim 4 achievable within the 5 year funding period, we will focus on MD analyses. Future research will analyze the predicted high-priority mutants in vitro and in vivo. We consider this proposal highly significant, because it implements and validates an efficient approach to understand the molecular mechanisms through which a therapeutically very important target class functions, interacts with small-molecule inhibitors and can become drug-resistant. If successful, our approach can be applied to other targets to identify drug-resistant mutants at the onset of a drug discovery project, enabling the structure-based rational design of molecules that inhibit wildtype and mutant kinases potently. This will aid the development of more selective, less side-effect and less drug-resistance prone therapeutics.

描述（由申请人提供）：激酶是第二大药物靶标家族，有10种批准的激酶抑制剂药物和50种化合物在临床试验中。蛋白质激酶结构域最常由癌症生成编码。其ATP结合G-Loops发生了几种癌症驾驶突变。 ABL抑制剂伊马替尼是慢性肌生型白血病的突破性治疗，但由于抗伊马替尼耐药的ABL激酶 - 瘤 - 瘤，尤其是在G-loop中，约有35％的患者复发。因此，由于使用激酶抑制剂药物治疗患者人群的增加，药物抗性可能成为一个主要的crincial问题。使用SRC家庭蛋白酪氨酸激酶LYN作为实验非常可触及的例子，我们建议实现和验证一种多学科方法，该方法首先使用分子动力学（MD）模拟来相对快速地识别影响催化和抑制剂相互作用的突变，并可能引起药物抵抗力（AIM 1）。我们接下来的方法分析了BA/F3细胞中鉴定出的LYN突变体对鉴定的Lyn突变体的活性，抑制作用以及抗性（AIM 2）或Lyn - / - 骨row（AIM 3），以识别那些在生理学上最相关的突变。在每个步骤中排除非信息突变体可以最大程度地减少实验努力，并最大程度地提高成功和成功的可能性。我们认为这种综合方法可以发现抗药性，从而导致激酶突变高度创新，因为它提供了重要的见解，通常只能在更长的时间段内和通过几个实验室的努力获得。这些研究跟进了我们最近发表的发现，58个真核激酶在其G-Loops上含有保守的静电盐桥，这对于G-环稳定，催化和ATP或ATP兼容抑制剂结合至关重要。 BCR-ABL中的盐桥破坏会导致伊马替尼的抗性。我们的初步数据表明，在31种激酶中，包括SRC，ABL，CK1和CK2-荷都在癌症中起重要作用，酸性盐桥锚也与保守的极性氨基或碱性氨基核或碱性 - acid-acid-acid-acid-acid-acid-side-side-side-side-side-side-side-side-side链相互作用。 To test the hypothesis that this "triad interaction-network" architecture is essential for G-loop function and inhibitor-interactions, and that its disruption can cause drug resistance, we will analyze the effects of mutationally modulating the different components of the variant G-loop-triad-configurations in the exemplary kinases Lyn (Aims 1-3), Abl, CK1(2 and CK2a1 （AIM 4）在5年的资金期间，我们将重点进行MD分析成功，我们的方法可以应用于其他靶标，以在药物发现项目开始时识别耐药突变体，从而实现了基于结构的合理设计，可抑制野生型和突变型激酶。这将有助于开发更具选择性，较少的副作用和耐药性易于治疗的疗法。