Developing a strategy to identify & validate pharmacodynamic kinase inhibitor biomarkers

制定策略来识别

• 批准号: 9197971
• 负责人: CHARLES J. BIEBERICH
• 金额: $ 20.2万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE COUNTY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-04 至 2018-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9197971
• 关键词: Address; Animal Model; Antibodies; Biological; Biological Assay; Biological Markers; Biopsy; Characteristics; Clinical; Clinical Trials; Development; Dose; Drug Targeting; Etiology; Failure; Gel; Goals; Industry; Investments; Knowledge; Label; MAPK1 gene; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Methods; Monitor; Mus; Outcome; Patients; Peptides; Pharmaceutical Preparations; Pharmacodynamics; Phosphorylation; Phosphotransferases; Physiological; Placebos; Play; Protein Kinase; Proteins; Proteome; Reaction; Reporting; Resolution; Role; Schedule; Site; Stable Isotope Labeling; System; Technology; Therapeutic; Time; Work; Xenograft procedure; biomarker development; biomarker panel; chemotherapy; cost effective; drug development; drug efficacy; in vivo; inhibitor/antagonist; innovation; inorganic phosphate; kinase inhibitor; liquid chromatography mass spectrometry; melanoma; new technology; novel strategies; oncology; pharmacodynamic biomarker; pre-clinical; protein profiling; public health relevance; response; response biomarker; small molecule; stable isotope; stoichiometry; success; targeted agent; targeted therapy trials; targeted treatment; tumor; tumor xenograft

 DESCRIPTION (provided by applicant): Small molecule kinase inhibitors comprise one of the most promising classes of oncology-directed therapies. Phenomenal clinical successes have already been achieved in a few cases, and intense efforts to broaden these successes are currently underway. More than 100 kinase inhibitors are currently in clinical trials, and hundreds of others are in the development pipeline. Unfortunately, it is highly likely that the vast majorit of these will fail, given that the current success rate for targeted oncology drugs is ~5%. A primary contributing factor to this unacceptably high failure rate is the paucity of pharmacodynamic biomarkers to assess drug efficacy at each stage of drug development. The dearth of kinase inhibitor biomarkers is the direct result of our more general lack of knowledge regarding the physiological targets of most protein kinases. In this application, we propose to develop and validate an innovative new approach for the identification of pharmacodynamic kinase inhibitor biomarkers. The current state-of-the-art in the industry is to employ anti-phosphosite antibodies to semi-quantitatively measure phosphorylation decreases in direct, or even indirect kinase substrates when the target kinase is inhibited. The choice of an appropriate biomarker is most often driven by anti-phosphosite antibody availability, rather than by pharmacodynamic characteristics of the responsive substrate. Here, we intend to vet a new paradigm that incorporates absolute quantification of phosphorylated and non-phosphorylated forms of inhibitor-responsive kinase substrates. We intend to profile protein phosphorylation stoichiometry in a kinase-specific manner, a goal that has not been achieved to date. This will be accomplished by incorporating stable isotope labeling into a reverse in-gel kinase assay, and analyzing tryptic peptides from ERK2 inhibitor-treated tumors using high resolution mass spectrometry. Quantification will be achieved using AQUA peptides as internal standards. By quantifying substrate responses at various intervals after drug administration, a panel of inhibitor-responsive substrates that accurately report on drug efficacy will be generated and validated. We anticipate that panels of biomarkers will outperform single biomarkers to determine optimal dose and scheduling of promising new kinase inhibitor therapies. The successful completion of this work will fulfill a pressing need to develop new technologies to reduce the failure rate of kinase inhibitors.

 描述（由申请人提供）：小分子激酶抑制剂是最有前途的肿瘤导向疗法之一，已经在一些案例中取得了显着的临床成功，并且目前正在进行 100 多个案例来扩大这些成功。激酶抑制剂目前正在进行临床试验，还有数百种其他抑制剂正在开发中，不幸的是，考虑到目前靶向肿瘤药物的成功率，其中绝大多数很可能会失败。导致这种不可接受的高失败率的一个主要因素是缺乏用于评估药物开发每个阶段的药物疗效的药效生物标志物。激酶抑制剂生物标志物的缺乏是我们普遍缺乏相关知识的直接结果。在此应用中，我们建议开发和验证一种创新的新方法来鉴定药效激酶抑制剂生物标志物。目前业界最先进的技术是采用抗磷酸位点抗体来识别。当靶激酶被抑制时，半定量测量直接甚至间接激酶底物的磷酸化减少。适当的生物标志物的选择通常由抗磷酸位点抗体的可用性驱动，而不是由响应底物的药效学特征驱动。我们打算审查一种新的范式，该范式结合了抑制剂响应激酶底物的磷酸化和非磷酸化形式的绝对定量。我们打算在蛋白质磷酸化化学计量中进行分析。激酶特异性方式，这一目标迄今为止尚未实现，这将通过将稳定同位素标记纳入反向凝胶内激酶测定中，并使用高分辨率质谱定量分析来自 ERK2 抑制剂治疗的肿瘤的胰蛋白酶肽来实现。我们预计，将使用 AQUA 肽作为内标来量化给药后不同时间间隔的底物反应，从而生成并验证一组能够准确报告药物功效的抑制剂反应底物。生物标志物组将优于单一生物标志物，以确定有前景的新型激酶抑制剂疗法的最佳剂量和时间安排。这项工作的成功完成将满足开发新技术以降低激酶抑制剂失败率的迫切需要。

项目成果

Quantifying Kinase-Specific Phosphorylation Stoichiometry Using Stable Isotope Labeling In a Reverse In-Gel Kinase Assay.

• DOI: 10.1021/acs.analchem.6b02599
• 发表时间: 2016-12-06
• 期刊: ANALYTICAL CHEMISTRY
• 影响因子: 7.4
• 作者: Li, Xiang;Cox, Jonathan T.;Huang, Weiliang;Kane, Maureen;Tang, Keqi;Bieberich, Charles J.
• 通讯作者: Bieberich, Charles J.