BCR Signaling in ABC DLBCL

目的：建立PCI-32765口服治疗复发性B细胞淋巴瘤患者的安全性和最大耐受剂量（MTD）。测定口服PCI-32765的药代动力学（PK）测量药效学参数，包括Btk的药物占用、靶酶和对B细胞功能生物标志物的影响。药代动力学和药效学样品将按照方案中概述的方式收集，并按照以下规定运送到Pharmacyclics， Inc.。组织样本不会被送到Pharmacyclics， Inc.。相反，组织样本将获得如下所述。在PCI-32765治疗开始前，所有患者都需要活检标本（治疗前活检）。预计大约一半的患者将有可接触的肿瘤组织，并将在PCI-32765之前进行重新活检。这些患者也将在初始PCI-32765剂量48小时后进行活检。对于其他患者，将分析以前活检的福尔马林固定和石蜡包埋（FFPE）样本。使用Affymetrix U133plus2.0阵列对所有病例进行基因表达谱分析。对于那些有可用的新鲜冷冻预处理活检样本的病例，将按照Staudt实验室的描述进行基因表达谱分析，并根据贝叶斯预测因子分配ABC或GCB亚组（Wright等）。中国科学院院刊，2003,31(1):1 - 3。对于这些患者，我们也将在治疗后48小时获得活检样本，并类似地分析基因表达。与预处理样品的比较将使我们能够评估PCI-32765抑制btk的能力。对于只有FFPE活检样本的患者，我们将使用一种新描述的方法，该方法可以使用FFPE活检的RNA来准确预测ABC与GCB DLBCL亚型（Williams等）。J Mol Diag 2010，已出版)。简单地说，使用Qiagen FFPE提取试剂盒从FFPE样品中提取RNA，然后使用Nugen试剂盒进行扩增，然后进行标准Affymetrix U133Plus2.0阵列分析。我们还将在可能的情况下进行肿瘤突变分析，包括但不限于CARD11、ITAM和A20。

Objectives: Establish the safety and the maximum tolerated dose (MTD) of orally administered PCI-32765 in patients with recurrent B cell lymphoma. Determine pharmacokinetics (PK) of orally administered PCI-32765 Measure pharmacodynamic parameters to include drug occupancy of Btk, the target enzyme, and effect on biological markers of B cell function Samples for pharmacokinetics and pharmacodynamics will be collected as outlined in the protocol and shipped to Pharmacyclics, Inc. as specified below. Tissue samples will not be sent to Pharmacyclics, Inc. Rather, tissue samples will be obtained as outlined below. Biopsy specimens will be required from all patients before initiation of PCI-32765 treatment (pre-treatment biopsies). It is anticipated that roughly half of the patients will have accessible tumor tissue and will be rebiopsied prior to PCI-32765. These patients will also be biopsied 48 hours after the initial PCI-32765 dose. For the other patients, formalin fixed and paraffin embedded (FFPE) samples from previous biopsies will be analyzed. Gene expression profiling using Affymetrix U133plus2.0 arrays will be performed for all cases. For those cases with available fresh frozen pre-treatment biopsy samples, geneexpression profiling will be performed as described by the Staudt laboratory, with assignment to ABC or GCB subgroups based on a Bayesian predictor (Wright et al. PNAS 2003, 100:9991). On these patients, we will also obtain biopsy samples 48 hours post-treatment and profile gene expression similarly. Comparison with the pre-treatment sample will allow us to assess the ability of PCI-32765 to inhibit btk. For patients with only an FFPE biopsy sample, we will use a newly described method that can use RNA from FFPE biopsies to accurately predict ABC vs. GCB DLBCL subtypes (Williams et al. J Mol Diag 2010, in press). Briefly, RNA is extracted from FFPE samples with a Qiagen FFPE extraction kit followed by amplification using a kit from Nugen, prior to standard Affymetrix U133Plus2.0 array analysis. We will also perform where possible analyses of tumor mutations which include but are not be restricted to CARD11, ITAM and A20.