Integrating Diagnostics with Therapeutic Strategies in Chronic Myeloid Leukemia

描述(申请人提供)：慢性粒细胞白血病(CML)提供了一种独特的疾病模型，在该模型中应用翻译方法来研究疾病进展、治疗反应和疾病复发的生物学。尽管BCR-ABL的功能已被很好地描述，但参与疾病进展的事件在很大程度上仍不清楚。在过去的10年里，酪氨酸激酶抑制剂如甲磺酸伊马替尼(IM)极大地改变了CML的治疗。然而，尽管大多数早期慢性期(CP)患者的反应良好，但约20%的CP患者IM治疗失败。这一数字对于晚期CP患者来说更高，而晚期疾病或对IM具有抵抗力的疾病的结果明显更差。因此，显然需要及早识别有疾病进展或治疗抵抗风险的患者。拟议的转化性研究依赖于确定患者的危险因素，然后在独立患者中进行验证，最终目标是将这些发现应用于临床环境中的患者护理。这些调查的成功取决于在足够多的患者中测试结果的能力。我们正在积极参与一些慢性粒细胞白血病的临床试验，包括SWOG S0325，它将标准剂量的IM与大剂量的IM和达沙替尼进行比较。在特定的目标1中，先前在微阵列研究中确定的与CML疾病进展和治疗耐药相关的候选诊断基因将在400个独立的患者样本中使用高通量定量RT-PCR(QPCR)进行验证。将通过对所有目前已知的miRNAs进行高通量QPCR分析，扩大研究范围，以确定与CML进展和治疗耐药性相关的microRNAs(MiRNAs)。在具体目标2中，这些诊断性研究将与接受酪氨酸激酶抑制剂(TKIs)治疗的患者的强化监测研究相结合。这些研究将包括早期检测和监测T315I的发育动力学，T315I是一种对目前批准的所有TKI都具有抗药性的突变。在保持最佳分子反应的患者中，还将使用一种新的纳米流体平台进行增强的BCR-ABL定量聚合酶链式反应监测，该平台通过将样本分割为多达10,000个独立的反应来提高灵敏度。T315I点突变研究的目的是确定是否可以使用早期T315I突变检测来指导治疗选择；对分子反应最佳的患者进行增强的BCR-ABL监测的目标是确定可能确定可以停止TKI治疗的患者的动力学特征。最后，在特定的目标3中，我们将使用基于慢病毒载体的基因沉默和表达技术来研究PRAME和GLI2在细胞系和原代细胞中在疾病进展和治疗耐药中的作用。一个主要目标将是确定参与进展和耐药的新途径，这些途径可以被现有的治疗方法或新的策略作为靶点。这些综合的生物学和临床研究未来的一个主要临床应用是确定可用于确定个别患者的基于风险的治疗管理的概况。公共卫生相关性：尽管甲磺酸伊马替尼(Gleevec，IM)在早期慢性粒细胞白血病(CML)患者中的治疗取得了成功，但对更晚期疾病或IM耐药疾病的患者的治疗结果仍然很差。我们已发表的和初步的数据使我们能够在临床试验样本中调查可能改变我们管理CML方式的几个重要问题：我们能否定义诊断反应预测因子；我们能否在治疗过程的早期确定泛耐药T315I突变；我们能否确定最佳应答者中的BCR-ABL mRNA模式，从而可能确定一组可能能够停止治疗的患者；以及通过更好地了解CML进展的生物学，我们能否确定治疗目标和耐药原因？这些发现将被用来创建基于风险的简档，最终将在临床环境中用于个别患者的量身定制治疗。

DESCRIPTION (provided by applicant): Chronic myeloid leukemia (CML) provides a unique disease model in which to apply a translational approach to the study of the biology of disease progression, therapeutic response, and disease relapse. Although the function of BCR-ABL is well described, events involved in disease progression remain largely unknown. Over the last 10 years, tyrosine kinase inhibitors such as imatinib mesylate (IM) have dramatically altered CML treatment. Yet despite the excellent responses in most early chronic phase (CP) patients, ~20% of CP patients fail IM therapy. This number is higher for late CP patients and outcomes for advanced disease or disease resistant to IM are significantly poorer. Thus, a clear need exists for the early identification of patients at risk for disease progression or therapeutic resistance. The proposed translational studies rely on the identification of risk factors in patients, followed by validation in independent patients, with the ultimate goal of applying these finding to patient care in the clinical setting. The success of these investigations relies on the ability to test the findings in a sufficiently large number of patients. We are actively participating in a number of clinical trials for CML, including SWOG S0325, which compares standard dose IM to high dose IM and to dasatinib. In Specific Aim 1 diagnostic gene expression candidates associated with CML disease progression and therapy resistance that were previously identified in microarray studies will be validated using high throughput quantitative RT-PCR (QPCR) in 400 independent patient samples. Investigations will be expanded to identify microRNAs (miRNAs) associated with CML progression and therapy resistance using high throughput QPCR profiling for all currently known miRNAs. In Specific Aim 2 these diagnostic studies will be integrated with enhanced monitoring studies in patients receiving tyrosine kinase inhibitors (TKIs). These investigations will include early detection and monitoring of the kinetics of T315I development, a mutation resistant to all currently approved TKIs. Enhanced QPCR monitoring of bcr-abl in patients maintaining the best molecular responses will also be performed using a novel nanofluidic platform that enhances sensitivity through sample partitioning into up to 10,000 independent reactions. The goal of the T315I point mutation studies is to determine whether early T315I mutation detection can be used to guide therapeutic choices; the goal of enhanced bcr-abl monitoring in the best molecular response patients is to determine a kinetic profile that may potentially identify patients in whom TKI therapy can be stopped. Lastly, in Specific Aim 3 we will examine PRAME and GLI2 for their role in disease progression and therapy resistance in cell lines and primary cells using lentiviral vector-based gene silencing and expression techniques. A main goal will be to identify new pathways involved in progression and resistance that can be targeted either by existing therapies or by novel strategies. A major future clinical application of these integrated biological and clinical studies is to identify profiles that can be used to determine risk-based treatment management in individual patients. PUBLIC HEALTH RELEVANCE: Despite the success of therapy with imatinib mesylate (Gleevec, IM) in early chronic phase chronic myeloid leukemia (CML) patients, treatment outcomes for patients with more advanced disease or IM resistant disease remain poor. Our published and preliminary data allow us to investigate in clinical trial samples several important questions that may change how we manage CML: can we define diagnostic predictors of response; can we identify the pan-resistant T315I mutation early in the treatment course; can we determine a bcr-abl mRNA pattern in the best responders that may identify a group of patients who may be able to stop therapy; and through a better understanding of the biology of CML progression can we identify targets for therapy and reasons for resistance? These findings will be used to create risk-based profiles that will ultimately be used in the clinical setting in individual patients to tailor therapy.