Role of CD28-mediated signaling in multiple myeloma

CD28介导的信号传导在多发性骨髓瘤中的作用

• 批准号: 144775016
• 负责人: Privatdozent Dr. Ingolf Berberich
• 金额: --
• 依托单位: Institut für Virologie und Immunbiologie
• 依托单位国家: 德国
• 项目类别: Clinical Research Units
• 财政年份: 2009
• 资助国家: 德国
• 起止时间: 2008-12-31 至 2012-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/144775016?language=en
• 关键词: Role; CD28; mediated; signaling; multiple

CD28 is a key co-stimulatory molecule expressed by naive and activated T cells, but is also found on short- and long-lived primary plasma cells (PC) and on mouse and human multiple myeloma (MM) cells, where its expression correlates with disease progression. Most published data suggest a role of CD28 in cell survival and sustained immunoglobulin (Ig) production, while others describe opposite effects.We found that ligation of CD28 with the superagonistic antibody TGN1412 was often not sufficient for activating signals (p-ERK, p-Akt, NFκB) in MM cells but required sub-signaling traces of phorbol-dibutyrate, suggesting that CD28 signaling depends on additional interactions with neighboring PC or other cells in BM niches.We have generated a mouse strain confining inactivation of CD28 to the B cell lineage by introducing an mb1Cre transgene into mice with a partially “floxed” CD28 gene. First data suggest diminished IgM and IgG production upon loss of CD28. However, no complete deletion of CD28 was achieved. To overcome this, we currently include breedings with CD19Cre, CD23Cre and AIDCre. We had also proposed to use B cell-specific CD28-deficient mice in a model of spontaneous MM. This had to be abandoned because compatibility with the use of Cre recombinase could not be established. We now turned to another model (iMyc mice), in which plasmacytoma are induced by pristane. We are currently crossing these mice to CD28fl/fl x mb1Cre mice. Here, incomplete deletion of CD28 will show if plasmacytoma preferably develop or expand when CD28 is present or not. In the meantime, we have exploited our established model of B lineage-specific ablation of the transcription factor C/EBPβ which shows impaired development of PC both in vitro and in vivo. Our results suggest that the two larger CEBPβ isoforms LAP* and LAP support survival and proliferation of human MM cell lines.For the next funding period, we propose to identify the putative natural co-factor required for effective CD28 signaling and the cell(s) expressing it. Likely co-stimuli provided by the interacting cells will be inhibited by blocking antibodies or mimicked by agonistic antibodies. A comprehensive analysis of the responding genes, will be performed by cDNA microarray analysis. With regard to in vivo studies, we will now apply the new inducible myeloma mouse model with lineage-specific and inducible ablation of CD28 expression to obtain answers about the role of CD28 for MM-development, survival and function. In addition, we will further dissect the role of B cell lineage-expressed CD28 (with different cre-deleters) for the humoral immune response and clarify the discrepancies with some reports from the literature.If CD28 is beneficial for MM survival, blocking antibodies which do not interfere with T cell activation could be a therapeutic approach. We will develop an antibody which recognizes CD28 with one arm and CD138 with the other one. If affinity is greatly boosted by dual binding, this bi-specific antibody would be a good candidate to test in our model systems.

CD28是一种关键的共刺激分子，由幼稚和活化的T细胞表达，但也存在于短寿命和长寿命的原代浆细胞（PC）以及小鼠和人多发性骨髓瘤（MM）细胞中，其表达与疾病进展相关。大多数已发表的数据表明CD28在细胞存活和持续免疫球蛋白（Ig）产生中的作用，而其他数据则描述了相反的作用。我们发现CD28与超激动抗体TGN1412的连接通常不足以激活MM细胞中的信号（p-ERK, p-Akt, NFκB），但需要phorbol2 - butyrate的亚信号通路，这表明CD28信号通路依赖于与邻近PC或BM龛中的其他细胞的额外相互作用。我们通过将mb1Cre转基因引入到部分“固定”CD28基因的小鼠中，产生了一种限制CD28失活的小鼠品系。第一项数据表明，失去CD28后，IgM和IgG的产生减少。然而，没有实现CD28的完全缺失。为了克服这一点，我们目前包括CD19Cre， CD23Cre和AIDCre的育种。我们也曾建议在自发性MM模型中使用B细胞特异性cd28缺陷小鼠。由于无法建立与使用Cre重组酶的相容性，因此不得不放弃。我们现在转向另一种模型（iMyc小鼠），其中浆细胞瘤是由嘌呤诱导的。我们目前正在将这些小鼠与CD28fl/fl x mb1Cre小鼠杂交。在这里，CD28的不完全缺失将显示当CD28存在时浆细胞瘤是否更容易发展或扩大。同时，我们已经建立了B谱系特异性消融转录因子C/EBPβ的模型，该模型在体外和体内都显示了PC的发育受损。我们的结果表明，两个较大的CEBPβ亚型LAP*和LAP支持人MM细胞株的存活和增殖。在下一个资助期，我们建议确定有效CD28信号传导和表达它的细胞所需的假定的天然辅助因子。相互作用细胞提供的可能的共刺激将被阻断抗体抑制或被激动抗体模仿。将通过cDNA微阵列分析对应答基因进行全面分析。在体内研究方面，我们现在将应用新的诱导性骨髓瘤小鼠模型，该模型具有谱系特异性和诱导性消融CD28表达，以获得关于CD28在mm发育，生存和功能中的作用的答案。此外，我们将进一步剖析B细胞谱系表达的CD28（具有不同的cred缺失）在体液免疫应答中的作用，并澄清与文献中一些报道的差异。如果CD28对MM存活有益，那么不干扰T细胞激活的阻断抗体可能是一种治疗方法。我们将开发一种单臂识别CD28，单臂识别CD138的抗体。如果双结合极大地提高了亲和力，那么这种双特异性抗体将是在我们的模型系统中测试的一个很好的候选抗体。