ANTI APOPTOTIC SIGNALING IN MYELOMA--BCL 2 AND NF KAPPA

骨髓瘤中的抗凋亡信号传导——BCL 2 和 NF KAPPA

• 批准号: 6514429
• 负责人: RENA R FEINMAN
• 金额: $ 21.2万
• 依托单位: UNIV OF MED/DENT OF NJ-NJ MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-04-01 至 2005-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6514429
• 关键词: BCL2 gene /protein; antisense nucleic acid; apoptosis; biological signal transduction; cell line; clinical research; dexamethasone; drug resistance; human subject; longitudinal human study; multiple myeloma; mutant; neoplasm /cancer chemotherapy; neoplasm /cancer pharmacology; nuclear factor kappa beta; oligonucleotides; posttranslational modifications

Understanding the molecular mechanisms by which myeloma cells evade signals leading to chemotherapy-induced apoptosis is the long term goal of this project. Glucocorticoids such as dexamethasone (Dex], all potent inducers of apoptosis are frequently used to treat multiple myeloma [MM], a clonal B cell-malignancy. Using standard doses of these agents, complete remissions are rare [5% of patients]. Drug resistance increases with prolonged treatment, suggesting primary resistance of tumor cells as a major obstacle to cure. This resistance maybe due to MM cell expression of moderate to high levels of the anti-apoptotic protein bcl-2. There is compelling evidence that bcl-2 imparts its anti-apoptotic signal by ducking and modulating the expression of genes critical for apoptosis, including transcription factors such as NF-kappaB. We hypothesize that chemoresistance in MM by bcl-2 requires activation of NF-kappaB. To investigate this hypothesis we will address the following specific aims: 1. Test whether persistent NF-kappaB activation observed in MM cells confers resistance in chemotherapy-induced apoptosis in MM cell lines. P50:p65 NF-kappaB subunits will be over-expressed in drug-sensitive MM cell lines to inactivated NF-kB. Antisense phosphorothiorate oligonucleotides for p50 and p65 will also be used to inhibit NF-kappaB. 2. Determine whether bcl-2 contributes to maintaining NF-kappaB activity through modulation of drug-mediated post-translational regulation of IkappaB proteins and/or p65 transactivation. Different modes of IkBalpha, Ikbeta and Ikepsilon protein regulation will be investigated: 1] basal and signal-induced IkappaB phosphorylation and degradation 2) IKK-alpha and -beta functions and 3) inhibitory properties of IkappaB proteins. For p65 transactivation: 1] a Gal4-BASED hybrid system will be used to test whether clinically relevant drugs and enforced bcl-2 interfere with the transactivation potential of p65 and 2] a bcl-2 deletion mutant that lacks the transmembrane domain will be expressed in MM cell lines to determine whether membrane-anchored bcl-2 augments p65 transactivation. 3. Investigate whether the inhibitory effects of Dex on NF-kappaB activity predict response to chemotherapy. Longitudinal studies will be conducted to determine whether patient responsiveness to glucocorticoid based therapy can predicted from their in vitro response to Dex in terms of NF-kappaB DNA binding, p65 immunostaining. There studies will hopefully provide deeper insight into the molecular basis of drug resistance in MM and should result in the identification of potential targets for therapeutic intervention.

了解该项目的长期目标是了解导致化疗引起的凋亡的分子机制。糖皮质激素（例如地塞米松（Dex）），所有有效的凋亡诱导剂经常用于治疗多发性骨髓瘤[mm]，一种克隆B细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 细胞 - 抗抑制作用，完全缓解的患者很少会受到抗药性的耐药性，这可能会导致较大的抑制作用，这表明该较大的电阻是较大的cymine抗性，而这些耐药性是cobiment的cymine抗性。高水平的抗凋亡蛋白Bcl-2有令人信服的证据表明，Bcl-2通过躲避和调节对凋亡至关重要的基因的表达，包括转录因子，包括NF-KAPPAB，我们通过BCL-2的范围来调查nf-kappab。在MM细胞中观察到的持续性NF-kappab激活是否在化学疗法诱导的MM细胞系中的抑制作用。 p50和p65的反义磷酸寡核苷酸也将用于抑制NF-kappab。 2。确定Bcl-2是否通过调节Ikappab蛋白和/或p65反式激活的药物介导的翻译后调节来维持NF-kappab活性。 Ikbalpha，Ikbeta和Ikepsilon蛋白调节的不同模式将进行：1]基础和信号诱导的Ikappab磷酸化和降解2）IKK-Alpha和-beta函数以及3）iKappab蛋白的抑制特性。对于p65反式活化：1]将使用基于GAL4的杂化系统来测试临床相关的药物和强制BCl-2是否会干扰p65和2] Bcl-2删除突变体的反式活化潜力，该突变体缺乏跨膜结构域在MM细胞线中表达的跨膜bclane-bcl-2 transactiv是否会在MM细胞线中表达。 3。研究DEX对NF-kappab活性的抑制作用是否预测了对化学疗法的反应。将进行纵向研究，以确定患者对基于糖皮质激素的治疗​​的反应是否可以从NF-kappab DNA结合，p65免疫染色的角度来预测其对DEX的体外反应。有望在那里提供对MM耐药性分子基础的更深入的了解，并应导致鉴定治疗干预的潜在靶标。