Cell-free DNA-Based Analysis for Diagnosis, Monitoring and Optimization of Therapy for Patients with Primary Central Nervous System Lymphomas

项目摘要 原发性中枢神经系统淋巴瘤是一种罕见的侵袭性淋巴结非霍奇金淋巴瘤 涉及中枢神经系统（CNS）而无全身性疾病。大约有1400例PCNSL病例 在美国，每年都有45岁至65岁的人被诊断出患有这种疾病，但发病率正在上升， 老年患者代表需要有效治疗且不良反应特征有限的人群[1，2]。标准 一线治疗包括大剂量甲氨蝶呤（HD-MTX）和自体干细胞移植作为巩固治疗 疗法然而，由于显著的神经毒性，并非所有患者都能耐受这些方案[3]。即使在那时， 三分之一的患者对一线治疗难治，高达60%的患者最终会复发[4]。预后 初治后难治或复发的患者预后较差，中位生存期仅为1年左右， 前期治疗[5]。由于长期结果不佳，迫切需要改进诊断和治疗 方式。进行临床试验以改善结果的一个相当大的障碍是缺乏可用的生物标志物， 可以可靠地用于诊断和监测疾病。因此，PCNSL的诊断通常需要神经外科手术。 活检是侵入性的，与感染、出血和神经损伤的风险相关。许多患者最初 接受腰椎穿刺（LP），以获取脑脊液（CSF），并通过细胞学和流式细胞术进行分析。这些 然而，这些方法缺乏灵敏度，需要大量（> 10 mL）CSF，并且在许多情况下是非诊断性的。 此外，作为疾病监测金标准的对比增强MRI不能检测早期复发或复发。 微小残留病（MRD）导致无法检测到化疗耐药性、完全缓解或复发。那里 不仅对PCNSL患者的更好治疗选择存在未满足的需求，而且对生物学生物标志物也存在未满足的需求， 有助于诊断和监测治疗反应。这些生物标记物在规划中将具有难以置信的重要性 并在PCNSL和相关疾病中进行未来的临床试验。 最近，几种靶向疗法，包括布鲁顿酪氨酸激酶抑制剂（例如，伊布替尼）以及 免疫调节剂（例如，来那度胺）已显示显著降低复发率[6-9]。分子 在PCNSL中，对这些新型靶向药物的治疗耐药性的基础在很大程度上是未知的，未表征的，并且是关键的 翻译研究领域。Safe-SeqS和Real-SeqS技术提供了检测和量化 CSF中的肿瘤DNA（CSF-tDNA），提供了更好地表征CNS疾病的机会。在本建议中，我们的目标是 开发稳健的Safe-SeqS和Real-SeqS检测试剂盒，用于检测肿瘤来源的循环游离DNA（cf-tDNA）， 提高我们诊断和基因分型PCNSL的能力，纵向监测疾病状态，并进行全外显子组 基因测序（WES），以确定分子特征，这些分子特征可以根据基因组序列识别患者进行靶向治疗。 响应的签名。从完成拟议目标中获得的知识将有助于制定 cf-tDNA作为PNCSL的第一个生物标志物，这是一种毁灭性恶性肿瘤急需的进展。

Project Summary Primary central nervous system lymphoma (PCNSL) is a rare and aggressive extranodal non-Hodgkin lymphoma that involves the central nervous system (CNS) without systemic disease. Approximately 1400 cases of PCNSL are diagnosed in the United States annually, classically occurring between the ages of 45 and 65, but the incidence is rising in older patients who represent a population in need of effective therapies with limited adverse effect profiles[1, 2]. Standard frontline therapy consists of high-dose methotrexate (HD-MTX) with autologous stem cell transplant as consolidation therapy. However, these regimens are not tolerated by all patients due to significant neurotoxicity[3]. Even then, about a third of patients are refractory to first-line treatment, and up to 60% of the patients will eventually relapse[4]. Prognosis for patients who are refractory to or relapse after initial therapy is poor, with median survival of only about 1-year failure of upfront therapy[5]. Due to poor long term outcomes, there is a desperate need for improved diagnostic and therapeutic modalities. A considerable hurdle in conducting clinical trials to improve outcomes is the lack of available biomarkers that can be used reliably to diagnose and monitor disease. As such, the diagnosis of PCNSL frequently requires neurosurgical biopsies that are invasive and associated with risk of infection, bleeding and neurological injury. Many patients initially undergo a lumbar puncture (LP) for cerebrospinal fluid (CSF) that is analyzed by cytology and flow cytometry. These approaches, however, lack sensitivity, require large quantities (> 10 mL) of CSF, and are non-diagnostic in many cases. Contrast-enhanced MRI, the gold-standard test of disease monitoring, furthermore, cannot detect early recurrence or minimal residual disease (MRD) leading to an inability to detect chemo-resistance, complete remission, or recurrence. There exists an unmet need not only for better treatment options for patients with PCNSL, but also for biological biomarkers to aid in diagnosis and monitoring of therapeutic response. Such biomarkers would be of incredible importance in planning and executing future clinical trials in PCNSL and related diseases. Recently, several targeted therapies, including Bruton tyrosine kinase inhibitors (e.g., ibrutinib) as well as immunomodulatory agents (e.g., lenalidomide) have shown to dramatically decrease recurrence rate [6-9]. The molecular basis for treatment resistance to these novel targeted agents in PCNSL is largely unknown, uncharacterized and a critical area of translational research. Safe-SeqS and Real-SeqS, technologies that provide an opportunity to detect and quantify tumor DNA in CSF (CSF-tDNA), provide an opportunity to better characterize CNS disease. In this proposal, we aim to develop robust Safe-SeqS and Real-SeqS assays for detection of tumor-derived circulating free DNA (cf-tDNA) which will improve our ability to diagnose and genotype PCNSL, to longitudinally monitor disease status, and to perform whole-exome sequencing (WES) to identify molecular characteristics that can identify patients for targeted therapies based on genomic signatures of response. The knowledge gained from the completion of the proposed Aims would enable the development of cf-tDNA as the first biomarker in PNCSL, a much needed advance for a devastating malignancy.