BCR Signaling in ABC DLBCL

目的：确定复发性B细胞淋巴瘤患者口服PCI-32765的安全性和最大耐受剂量（MTD）。测定PCI-32765口服给药的药代动力学（PK）测量药效学参数，包括Btk（靶酶）的药物占有率和对B细胞功能生物标志物的影响。将按照方案中的概述收集药代动力学和药效学样品，并运送至Pharmacyclics，Inc.。如下所述。组织样本将不会发送至Pharmacyclics，Inc.。相反，将按照下文所述获得组织样本。在开始PCI-32765治疗前，需要所有患者的活检标本（治疗前活检）。预计大约一半的患者将有可触及的肿瘤组织，并将在PCI-32765之前进行重新活检。这些患者还将在初始PCI-32765给药后48小时进行活检。对于其他患者，将分析既往活检的福尔马林固定和石蜡包埋（FFPE）样本。将对所有病例进行使用Affytelium U133plus2.0阵列的基因表达谱分析。对于具有可用的新鲜冷冻治疗前活检样品的那些病例，将如施陶德德实验室所述进行基因表达谱分析，并基于贝叶斯预测因子分配至ABC或GCB亚组（Wright et al. PNAS 2003，100：9991）。对于这些患者，我们还将在治疗后48小时获得活检样本，并以类似方式分析基因表达。与治疗前样品的比较将使我们能够评估PCI-32765抑制btk的能力。对于仅有FFPE活检样本的患者，我们将使用一种新描述的方法，该方法可以使用来自FFPE活检样本的RNA准确预测ABC与GCB DLBCL亚型（威廉姆斯等人，J Mol Diag 2010，出版中）。简而言之，在标准Affytron U133Plus2.0阵列分析之前，用Qiagen FFPE提取试剂盒从FFPE样品中提取RNA，然后使用来自Nugen的试剂盒扩增。我们还将在可能的情况下对肿瘤突变进行分析，包括但不限于CARD 11、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.