Design, Develop, Validate, and Implement Biomarkers for Clinical Investigations

背景：靶向抗癌治疗的发展需要 证明研究制剂对预期靶点和途径的影响 以及发现与反应相关的标记。因此，我们 致力于检测试剂对途径特异性生物标志物的影响， 在相关研究中实施反应生物标志物。I.技术，检测设计， 开发和验证。我们开发、验证和实施临床标本检测 使用基于电化学发光（ECL）的免疫测定。这是最敏感的， 定量免疫分析技术平台。ECL平台非常适合于此 这是一项持续的任务，因为它提供了高度的灵活性，稳定性和可靠性。是 能够多重分析，以确定总蛋白和磷蛋白的水平， 使用有限数量的临床样本的单个测定孔。因为临床样本可能 差异很大，将这些样品标准化超过总蛋白浓度的能力 对于患者标本生成统计学显著数据至关重要。我们有 成功地利用免疫分析技术检测ERK、AKT 通路、mTOR抑制、血管生成因子以及临床中的多种细胞因子 国家癌症研究所的规程我们目前是NCI唯一一个提供服务和协作的实验室 在这些平台上进行分析。我们不断挑战极限， 能够满足临床试验中未满足需求的检测。二.当前和最近 已完成的生物标志物研究。目前，我们在NCI-CCR参与了10项临床方案。 对于其中许多临床试验，我们帮助设计、开发、验证和实施 用于相关分析研究的定制生物标志物测定。以下是一些 例如：1）索拉非尼在雄激素非依赖性前列腺中的II期临床试验 癌症（PI，William Dahut），我们的实验室进行了试验验证， 药物索拉非尼，并使用骨髓活检标本进行测定，以确定其 对靶向MAP激酶途径的作用（DahutWL等，Clin.CancerRes.14：209 - 14， 2008年）。2)另一项研究联合靶向治疗的临床试验 卵巢癌（PI，Elise Kohn），我们的小组开发了检测方法，并进行了分析， 血管生成因子血管内皮生长因子（VEGF）和细胞因子，包括 白细胞介素-6（IL6）和白细胞介素-8（IL8）从患者血浆中获得， 药物试验期（Azad NS等人，J. Clin. Oncol. 26：3709 - 14，2008）。3)在另一 雷帕霉素试验（PI，Chand卡纳）中，我们测量了S6 RP的相对变化 来自骨肉瘤活检和外周血单核细胞（PBMC）的信号传导 在给予雷帕霉素之后。我们的数据显示， PBMC中p-S6RP的减少。p/t-S6RP比值从给药前25%变为给药后0.5% p 0.001，样本从第2天到研究结束。<我们的结果 进一步表明，在肿瘤活检中存在高度变异。的统计学 雷帕霉素对S6RP途径的显著作用只能通过p/t-S6RP比率来证明 使用我们的双重测定能够定量测量p-S6RP和t-S6RP， （已提交手稿）。三.支持药物的临床前研究 发展我们参与了广泛的临床前研究，以支持 NCI的临床研究。一些例子是：1）发现用于以下的预测生物标志物： 与Lee Helman博士合作的抗IGF1R抗体治疗横纹肌肉瘤。（二） 抗IGF1R治疗胸腺癌的可行性研究与Giuseppe Giaccone博士。第三章 TORC1/2抑制剂Palomid 529与放射联合治疗对头部和颈部的影响 与DCTD的Stephen Yoo博士合作治疗颈部癌症。4)的研制与评价 高灵敏度测定作为HIF 1a抑制剂生物标志物的HIF 1a测定。总之，生物标志物 药效学生物标志物和预测性生物标志物的分析是 早期癌症药物开发。我们在分析开发方面具有独特的能力， 我们积极参与NCI的临床试验。

Background: The development of targeted anti-cancer therapies requires the demonstration of the effects of the investigation agents on the intended target and pathway and the discovery of the markers that can be associated with the response. Thus, we are engaged in detecting the effects of agents on pathway-specific biomarkers and in discovering response biomarkers to be implemented in correlative studies. I. Technology, Assay Design, Development, and Validation. We develop, validate, and implement assays for clinical specimens using electrochemiluminescence (ECL)-based immunoassays. This is the most sensitive and quantitative immunoassay technology platform today. The ECL platform is well suited for this ongoing task because it offers a high degree of flexibility, stability and reliability. It is capable of multiplex analysis to determine the levels of total and phospho-proteins in a single assay well using a limited amount of clinical specimens. Because clinical samples may vary dramatically, the ability to normalize these samples beyond total protein concentration is critical in generating statistically significant data with patient specimens. We have successfully utilized the technology for immunoassays that detect changes in ERK, AKT pathways, mTOR inhibition, angiogenic factors, as well as a variety of cytokines in clinical protocols at NCI. We are currently the only lab at NCI provides services and collaboration with assays on these platform. We are constantly pushing the envelope to come up with new assays capable of meeting the unmet needs in clinical trials. II. Current and Recently Completed Biomarker Studies. Currently, we are engaged with 10 clinical protocols at NCI-CCR. For many of these clinical trials, we helped to design, develop, validate, and implement customized biomarker assays for correlative analytical studies. The following are some examples: 1) For a phase II clinical trial of sorafenib in androgen-independent prostate cancer (PI, William Dahut), our laboratory performed assay validation with the investigational agent sorafenib and implemented assays with bone marrow biopsy specimens to determine its effects on the targeted MAP kinase pathway (Dahut WL, et al. Clin. Cancer Res. 14: 209-14, 2008). 2) For another clinical trial that investigated a combination targeted therapy against ovarian cancer (PI, Elise Kohn), our group developed assays and performed the analysis of angiogenic factor vascular endothelial growth factor (VEGF) and cytokines, including interleukin-6 (IL6) and interleukin-8 (IL8) from plasma obtained from the patients throughout the drug trial period (Azad NS, et al. J. Clin. Oncol. 26: 3709-14, 2008). 3) In another rapamycin trial (PI, Chand Khanna), we measured the changes in the relative changes of S6RP signaling from both osteosarcoma biopsies and peripheral blood mononuclear cells (PBMC) following the administration of rapamycin. Our data showed a highly statistically significant reduction of p-S6RP in PBMC. The p/t-S6RP ratio changed from 25% pre-drug to 0.5% post-drug with p<0.001, with samples obtained from day 2 to the end of the study. Our results further showed that there is a high degree of variation in tumor biopsies. The statistically significant effect of rapamycin on the S6RP pathway can only be demonstrated as p/t-S6RP ratio using our duplex assay capable of quantitatively measuring both p-S6RP and t-S6RP in the same assay well (manuscript submitted). III. Preclinical investigations in support of drug development. We are engaged with a wide range of preclinical investigations in support of clinical studies at NCI. Some examples are: 1) Discovering the predictive biomarker for anti-IGF1R antibodies against rhabdomyosarcoma in collaboration with Dr. Lee Helman. 2) Feasibility studies for anti-IGF1R therapy in thymic cancers with Dr. Giuseppe Giaccone. 3) Evaluation of TORC1/2 inhibitor Palomid 529 in combination with radiation against head and neck cancers in collaboration with Dr. Stephen Yoo of DCTD. 4) Development and evaluation of highly sensitive assay HIF1a assay as a biomarker for HIF1a inhibitors. In summary, biomarker analysis for pharmacodynamic biomarkers and predictive biomarkers is an essential part of early stage cancer drug development. We have unique capabilities in assay development, validation and implementation, and we actively engage clinical trials at NCI.