Biosensor technology to monitor leukemia-related kinase activity in patient cells

描述(由申请人提供)：即使最近开发了像格列卫这样的重磅炸弹酶抑制剂药物，白血病仍然是致命癌症的高位。其中一个原因是患者对抑制剂药物的反应是通过间接手段(血液学缓解、细胞遗传学反应、信使核糖核酸表达水平)监测的。我们正在结合两种新兴的方法：我们基于多肽的靶向细胞内激酶传感器和多反应监测(MRM)质谱仪，以开发一种激酶分析技术，该技术将从患者材料中提供有关激酶抑制的直接信息。该策略还将兼容同时分析多个激酶，因此可以解决磷酸化信号转导的系统级生物学问题。在这项初步研究中，我们将检查各种参数，以确定最佳传感器浓度、细胞数量、分析方案和统计分析，以从患者材料中获得有意义的激酶活性数据。我们还将优化样本处理方案，以便在进行激酶检测之前收集、处理和存储患者材料。在成功演示和优化这项技术后，我们将准备通过使用R33机制的额外技术开发和/或通过假设驱动机制进一步研究白血病中的信号生物学来扩大这一项目。我们预计，我们的技术将为检测原发患者细胞中的激酶活性提供前所未有的灵敏度和细节水平，并有一天可能成为相对“实时”监测抑制剂药物效果的工具，使医生有机会调整剂量，以使他们的患者获得最大限度的激酶抑制剂治疗的积极结果。 与公共卫生相关：我们正在开发监测白血病药物对激酶抑制的技术。我们的分析结合了两项新兴技术，潜在地提供了有关患者药物反应的信息，医生可以利用这些信息来调整剂量和改善治疗。

DESCRIPTION (provided by applicant): Even with the recent development of blockbuster kinase inhibitor drugs like Gleevec, leukemias are still high on the list of deadly cancers. One reason for this is that patient response to inhibitor drugs is monitored by in- direct means (hematological remission, cytogenetic response, mRNA expression levels). We are combining two emerging methods: our peptide-based targeted intracellular kinase sensors and Multiple Reaction Monitoring (MRM) mass spectrometry to develop a kinase assay technology that will provide direct information about kinase inhibition from patient material. The strategy will also be compatible with simultaneous analysis of more than one kinase at a time, so the systems-level biology of phosphorylation signaling can be addressed. In this pilot study, we will examine various parameters to determine the optimal sensor concentration, cell number, analytical protocols and statistical analysis to obtain meaningful kinase activity data from patient material. We will also optimize the sample handling protocols for collecting, processing and storing the patient material prior to the kinase assay. Upon successful demonstration and optimization of this technique, we will be poised to expand this project through additional technological development using the R33 mechanism, and/or further study of signaling biology in leukemia through a hypothesis-driven mechanism. We anticipate that our technology will provide unprecedented levels of sensitivity and detail for examining kinase activity in primary patient cells, and may someday become a tool for monitoring inhibitor drug effects in relatively 'real-time,' giving physicians the opportunity to adjust dosage to maximize the positive outcomes of kinase inhibitor treatment for their patients. PUBLIC HEALTH RELEVANCE: We are developing technologies to monitor the inhibition of kinases by drugs in leukemia. Our assay combines two emerging technologies to potentially provide information about drug response in patients that physicians could use to adjust dosages and improve treatment.