Digital PCR quantification of BCR-ABL for CML diagnosis and monitoring in a LMICs setting

项目摘要 我们建议开发一种低资源设置(LRS)数字聚合酶链式反应(DPCR)仪来检测bcr-abl 从小血样中提取转录本以识别符合条件的慢性粒细胞白血病(CML)患者 低收入和中等收入国家(LMIC)的酪氨酸激酶抑制剂(TKI)治疗。尽管取得了成功， 靶向TKI Glivec(格列卫、伊马替尼)在发达国家治疗CML中的作用，世界上大多数 慢性粒细胞白血病患者居住在LMIC中，获得诊断检测和TKI治疗的机会有限。《欢乐合唱团》 国际患者援助计划(GIPAP)是全球范围内最全面、影响最深远的计划之一 由诺华公司(Glivec的制造商)与MAX合作设计的癌症获取计划 基金会(TMF)通过以下途径促进Glivec(伊马替尼)的获得和直接分发给符合条件的患者 他们的提供者。实施GIPAP计划的主要挑战之一是缺乏诊断 许多GIPAP国家的能力对于选择接受TKI治疗的患者至关重要，因为只有 被正确诊断为费城染色体阳性CML(Ph+CML)的患者有资格 参与GIPAP计划。我们假设一台检测bcr-abl的LRS dPCR仪器 高度敏感的文字记录将使更多具有LMICs能力的医疗机构能够诊断 Ph+CML和参与GIPAP计划，以及监测正在接受TKI治疗的患者。 我们将采用我们实验室最新开发的技术来开发LRS dPCR仪 包括一种基于自数字化(SD)微流控芯片的新型数字核酸扩增平台， 它将水样分割成数万纳升的体积，适合用于聚合酶链式反应仪器 对于使用离心力的有效样品加载和使用光盘(OD，例如CD，DVD， 蓝光)样式的读卡器，与LMIC设置高度兼容。我们建议利用这些 创新技术，创造一种符合或超过灵敏度和特异度的LRS dPCR仪器 最先进的RT-PCRbcr-abl融合转录本分析，只需最少的样品制备和 无需运行即可运行标准。这样的工具将对改善CML产生重大影响 患者在LMICs中的存活率。

Project Abstract We propose to develop a low resource setting (LRS) digital PCR (dPCR) instrument to detect BCR-ABL transcripts from small blood samples to identify chronic myeloid leukemia (CML) patients who are eligible for tyrosine kinase inhibitor (TKI) therapy in low- and middle- income countries (LMICs). Despite the success of the targeted TKI Glivec (Gleevec, imatinib) in treating CML in developed countries, the majority of the world's CML patients reside in LMICs with limited access to diagnostic testing and TKI treatment. The Glivec International Patient Assistance Program (GIPAP) is one of the most comprehensive and far-reaching global cancer access programs designed by the Novartis (the manufacturer of Glivec) in partnership with the Max Foundation (TMF) to facilitate access to and distribution of Glivec (imatinib) directly to eligible patients through their providers. One of the major challenges of the implementation of GIPAP program is the lack of diagnostic capabilities in many GIPAP countries which are essential for selecting patients for the TKI therapy, since only patients who are properly diagnosed with Philadelphia chromosome positive CML (Ph+ CML) are eligible to participate in the GIPAP program. We hypothesize that a LRS dPCR instrument to detect the BCR-ABL transcript in a highly sensitive manner will enable more health care institutes in LMICs capabilities to diagnose Ph+ CML and participate in the GIPAP program, as well as monitor patients who are being treated with TKI. We will develop the LRS dPCR instrument by adapting technologies recently developed in our laboratories including a novel digital nucleic acid amplification platform based on a self-digitization (SD) microfluidic chip, which partitions an aqueous sample into tens of thousands of nanoliter volumes suitable for PCR, instruments for efficient sample loading using centrifugal force and signal detection using optical disc (OD, e.g. CD, DVD, Blu-Ray)-styled readers which are highly compatible to the LMICs setting. We propose to leverage these innovative technologies to create a LRS dPCR instrument matching or exceeding the sensitivity and specificity of the current state of the art RT-PCR BCR-ABL fusion transcript assay, with minimal sample preparation and without the need of run to run standards. Such an instrument will have significant impact on improving CML patients' survival in LMICs.