铜螯合剂由抑制SOD1的活性调控胞内氧化还原信号转导

Cu/Zn superoxide dismutase (SOD1) maintains the homeostasis of superoxide anion and peroxide hydrogen by catalyzing the dismutation of superoxide to peroxide hydrogen. NADPH oxidase (Nox) acts as one of the main superoxide sources. Upon the stimulation by the binding of cytokines to their receptors on membrane, Nox is activated, leading to the formation of superoxide at a high level.Thereby, SOD1 results in a significant increase in the concentration of peroxide hydrogen. Peroxide hydrogen at the high levels can inactivated signaling proteins by reversibly oxidazing the cysteine residues in their active sites, resulting in activation of the aberrant redox signaling network or dysregulation of the redox signaling network. This can cause not only the cellular responses such as hypertrophy, proliferation, and migration, but also angiogensis, leading to tumorigensis. Moreover, the aberrant redox signaling may cause inflammation and damage motor neurons, resulting in amyotrophic lateral sclerosis (ALS). Therfore, SOD1 has become a target for designing anticancer and anti-ALS drugs. Thioflavin T (ThT) is a fluorescent reagent sensetive to beta-sheet proteins and their aggregates, and the Cu bio-probes have a strong affinity for the intracellular Cu. Through rational combination of the recognition property of ThT for beta-sheet protein structures and the chelating ability of the intracellular Cu probes, a specific and strong chelating reagent for the Cu in SOD1 can be designed. The chelating reagent inhibits SOD1 by removing the Cu in SOD1, which can notably alter the relative concentrations of intracellular superoxide and peroxide hydrogen. This alteration in ROS levels may lead to selective apoptosis or necrosis of the cells. Therefore, the affected redox signaling pathways can be understood at molecular levels. The study is valuable in designing the drugs that can not only inhibit the cell proliferation and angiogensis, but also attenuate the motor neuron damage via rationally regulating the redox signaling network.

铜锌超氧化物歧化酶(SOD1)催化超氧阴离子歧化为过氧化氢，维持胞内超氧和过氧化氢的内稳态浓度。NADPH氧化酶作为超氧的主要源之一，受细胞因子与其膜表面受体结合的刺激而活化将产生高浓度超氧，SOD1则导致胞内过氧化氢浓度显著上升。高浓度过氧化氢由可逆氧化活性部位的半胱氨酸残基使信号蛋白失活，激活胞内异常的氧化还原信号转导网络，或使正常的氧还信号网络失调。这既能导致细胞过度增殖，新血管生成，引起癌变；又能引起严重的炎症，损伤运动神经元，导致脊髓侧索硬化症(ALS)。因此，SOD1成了设计抗癌和抗ALS药物的重要靶标。合理组合beta折叠蛋白识别试剂和灵敏的胞内铜成像试剂，设计特异的螯合剂由去除SOD1中的铜使其失活，改变胞内超氧和过氧化氢的相对水平，调控氧还信号途径，由此选择性地导致细胞凋亡或死亡。这有助于设计由合理调控氧还信号网络，抑制细胞过度增殖和血管生成、减轻运动神经元损伤的药物。

围绕项目申请书提出的主题：设计高效铜螯合剂由特异抑制SOD1的活性调控胞内氧化还原信号转导，发展以SOD1为靶的新型抗癌药物。我们首先设计合成了几个系列的对SOD1具有显著抑制作用的螯合剂，经筛选获得的代表性螯合剂较文献报导的螯合剂如二乙基二硫代氨基甲酸钠（DDC）和四硫代钼酸盐类（MoS42-，TM）等具有抑制效率高，跨膜容易，特异性好等优点。其次我们就抑制剂对胞内ERK、PI3K信号通路及细胞凋亡的影响等方面展开了系统研究，发现螯合剂通过嵌入SOD1底物通道螯合铜抑制SOD1活性，由抑制SOD1准确调控胞内ROS（H2O2；O2•−）相对浓度，ROS浓度变化对ERK、PI3K信号通路及癌细胞凋亡有显著影响。这些结果对我们理解ROS信号通路中各种激酶的激活机制乃至多种病理生理过程具有十分重要的意义，如从信号转导角度揭示SOD1导致ALS的化学基础，发展有效治疗ALS的新途径；发现SOD1活性变化与某些癌症发生、发展之间的关系，为设计以SOD1为靶的新型抗癌药物打下基础。