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Studies on abnormalities of membrane receptor signaling in malignant lymphoma cells as a basis of diagnosis and treatment

恶性淋巴瘤细胞膜受体信号异常作为诊断和治疗依据的研究

基本信息

批准号：
16390103
负责人：
WATANABE Toshiki
金额：
$ 9.54万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16390103/
关键词：
cancer signal transduction lymphoma diagnosis molecular pathophysiology CD30 NF-kB TRAF protein HTLV-1 ATL DNA microarray expression profile Hodgkin lymphoma

项目摘要

(1)Analysis of CD30 promoter activation :We demonstrated that the constitutive overexpression CD30 activate ERK1/2 MAPK pathway, and induces JunB expression, which is a common characteristic of neoplastic. Activation of NF-kB results from "ligand-independent activation" of overexpressed CD30 on the Hodgkin/Reed- Sternberg (H-RS) cells, and that inhibition of CD30 signaling by transduction of a dominant negative decoy CD30 that lacks the cytoplasmic region induced apoptotic cell death of H-RS cells (Horie et al., Oncogene 21, 2493, 2002). We showed that cytoplasmic aggregation of TRAF2 and TRAF5 proteins represents constant signaling of CD30 ( Horie et al., Am J Pathol 160, 1647, 2002). We demonstrated that the AP-1 binding sequence located near the microsatellite counteracts the suppressive effects of the microsatellite on the CD30 promoter activity (Watanabe et al., Am J Pathol 163, 633, 2003). We showed that p80NPM-ALK inhibits ligand-independent signal transduction by overexpressed … More CD30 in ALCL cells though sequestration of TRAF proteins in the complex of NPM-ALK and wild type NPM (Horie et al., Cancer Cell 5, 353, 2004).(2)Expression profile analysis of primary ATL cells :We characterized the expression profile of primary ATL cells using the synthetic DNA array system and samples of 16 ATL cases and 11 healthy volunteers. The results revealed 108 overexpressed genes and 250 downregulated genes. Based on the results, we are now preparing a detection system of ATL cells in the PBMC using multiple real-time PCR analysis of samples. A pilot study clearly detected ATL cells in a sample derived from a carrier considered to be an asymptomatic carrier, which was confirmed by monoclonal pattern in subsequent Southern blot analysis.(3)Studies on the target genes of NF-kB :Expression profile analyses were done by CodeLink system using cell lines with constitutive activation of NF-kB and treatment by DHMEQ. The results revealed more than 1,000 genes that are up- or down-regulated more than two-fold by DHMEQ treatment. Up-regulated genes included these involved in proapoptotic function, and down-regulated genes included those involved in anti-apoptotic function. Furthermore, many genes were revealed to be target genes of NF-kB signaling (a part of these data were used in the manuscript of BLOOD 106, 2642, 2005).(4)Basic studies on the molecular targeted therapy by NF-kB inhibition using DHMEQ :Anti-tumor activity of DHMEQ against ATL, CLL and Hodgkin's lymphoma was tested by in vitro experiments and in vivo xenograft model using a SCID mice system. As to ATL, DHMEQ was shown to be able to purge carrier's PBMC of HTLV-1-infected but not transformed cells (BLOOD 106, 2642, 2005). DHMEQ was also effective against CLL cells, and showed synergistic effect with fludarabine when used in combination (Leukemia 20, 800, 2006). DHMEQ showed anti-tumor activity against H-RS cells in vitro and in vivo (Cancer Res, submitted). Less

(1)CD30启动子激活分析：我们证明组成型过表达CD30激活ERK1/2 MAPK通路，并诱导JunB表达，这是肿瘤的共同特征。 NF-kB 的激活是由 Hodgkin/Reed-Sternberg (H-RS) 细胞上过表达的 CD30 的“配体依赖性激活”引起的，并且通过转导缺乏细胞质区域的显性失活诱饵 CD30 来抑制 CD30 信号传导，诱导 H-RS 细胞凋亡（Horie 等人，Oncogene 21, 2493, 2002）。我们表明 TRAF2 和 TRAF5 蛋白的细胞质聚集代表 CD30 的恒定信号传导（Horie 等人，Am J Pathol 160, 1647, 2002）。我们证明位于微卫星附近的 AP-1 结合序列抵消了微卫星对 CD30 启动子活性的抑制作用（Watanabe 等人，Am J Pathol 163, 633, 2003）。我们发现，p80NPM-ALK 通过在 NPM-ALK 和野生型 NPM 复合物中隔离 TRAF 蛋白，在 ALCL 细胞中过表达 CD30，从而抑制配体依赖性信号转导（Horie 等人，Cancer Cell 5, 353, 2004）。 (2) 原代 ATL 细胞的表达谱分析：我们使用合成 DNA 阵列系统和样品来表征原代 ATL 细胞的表达谱。 16 名 ATL 病例和 11 名健康志愿者。结果显示有 108 个过表达基因和 250 个下调基因。基于这些结果，我们现在正在准备使用对样品进行多次实时 PCR 分析的 PBMC 中 ATL 细胞的检测系统。一项初步研究在来自被认为是无症状携带者的样本中清楚地检测到了 ATL 细胞，这在随后的 Southern blot 分析中通过单克隆模式得到了证实。 (3)NF-kB 靶基因的研究：使用 NF-kB 组成型激活并经过 DHMEQ 处理的细胞系，通过 CodeLink 系统进行表达谱分析。结果显示，超过 1,000 个基因在 DHMEQ 治疗后上调或下调了两倍以上。上调基因包括那些参与促凋亡功能的基因，下调基因包括那些参与抗凋亡功能的基因。此外，许多基因被揭示为NF-kB信号传导的靶基因（其中部分数据被用于BLOOD 106、2642、2005的手稿中）。（4）利用DHMEQ抑制NF-kB分子靶向治疗的基础研究：通过体外实验测试了DHMEQ对ATL、CLL和霍奇金淋巴瘤的抗肿瘤活性。以及使用 SCID 小鼠系统的体内异种移植模型。至于ATL，DHMEQ被证明能够清除载体PBMC中HTLV-1感染的但不能转化的细胞(BLOOD 106, 2642, 2005)。 DHMEQ 对 CLL 细胞也有效，并且与氟达拉滨联合使用时显示出协同作用（Leukemia 20, 800, 2006）。 DHMEQ 在体外和体内均表现出针对 H-RS 细胞的抗肿瘤活性（Cancer Res，已提交）。较少的