质谱在金属抗癌药物与蛋白质相互作用研究中的应用

研究细胞毒性金属抗肿瘤药物与蛋白质的相互作用,以及这种相互作用对药物的细胞摄入、转运、代谢和生物利用度的影响,对金属抗癌药物的结构设计和优化,提高药物的抗癌活性,降低毒副作用具有重要意义。基于软电离技术的电喷雾质谱和基质辅助激光解析电离质谱能够在分析检测过程中很好的保留金属抗癌药物与蛋白质的共价(配位)结合,获得药物与蛋白质结合位点的信息。同时,质谱分析还具有灵敏度高,所需样品量少,耗时短以及适用于分析复杂生物样品等优点,已成为研究金属抗癌药物与蛋白质相互作用最强有力的工具,在为药物发现提供大量化学、生物信息的同时,也极大地促进了质谱技术自身的发展。本文将结合我们在金属抗癌药物相互作用组学研究中取得的最新进展,系统地总结、评述Bottom-up和Top-down质谱分析方法在铂、钌类金属抗癌药物与蛋白质相互作用研究中的发展动态,并分析这一前沿交叉领域未来的发展趋势。     

抗肿瘤药物顺铂和血清白蛋白的相互作用研究

Cisplatin is a widely used anticancer agent to treat solid tumours such as ovarian, testicular cancers. The interaction of platinum drugs with human albumin is an important issue that contributes to understanding the transport process of platinum drugs, cell uptake and their side-effects. Here, we applied a bottom-up proteomic approach to study the interaction of cisplatin with recombinant human albumin(rHA). The cisplatin-rHA complexes were prepared under physiological conditions in vitro at different cisplatin/rHA molar ratios for different incubation times. LC/MS analysis enables to identify four platinated peptides and further MS/MS analysis characterizes the platinum binding sites. The results show that cisplatin binds to His67, His128, His247 and Met298 residues in albumin involving an intramolecular crosslinking of His67 to His247 by platinum. Besides, cisplatin is shown to induce the cleavage of the disulfide bond Cys124-Cys169 of albumin and bind to the thiol in the reduced Cys124 residue.     

有机金属钌抗癌药物与G-四链体DNA的相互作用研究

Guanine-rich sequence TTAGGG can form G-quadruplex(G4) at the end of human telomeres, which protect chromosomal ends from unwanted recombination and degradation and inhibit the activity of telomerase enzyme, The telomerase was shown to be active in 85%-90% of human cancer cells, but inactive in healthy and somatic cells. DNA is the potential target of organometallic ruthenium(II) anticancer complexes. It is of great importance to study the interaction of ruthenium complex with the senior structural DNA G4. The present work focused on investigating the interactions between organometallic ruthenium complexes and G-quadruplex. The pilot studies show that NH4^＋ can stabilize the G4 structure to reduce the binding of biphenyl ruthenium complexes to guanine bases, but the inhibitory effects disappeared as the concentration of ruthenium complexes increases. This result suggests that the coordination of ruthenium complexes may distort and even unwind the G-quadruplex so that further coordination of ruthenium to the linear DNA fragment occurred.     

MALDI-TOFMS快速筛选蛋白激酶抑制剂的方法初探

A MALDI-TOF mass spectrometric method for rapid screening of protein tyrosine kinase(PTK) inhibitors was developed. Due to the low abundance and low ionization efficiency of phosphopeptides, a separation and enrichment process was carried out by using a house-made TiO2 nanoparticle-coated capillary column prior to the MS analysis. With a synthetic phosphopeptide, of which the sequence is similar to that of the substrate peptide, as the internal standard, the ratios of signal intensities of the standard to the substrate are linear to the molar ratios of the two phosphopeptides over the range of 0.3 to 3. The proposed method will be applied to screen PTK inhibitors synthesized in our laboratory.     

铂基抗癌药物与蛋白质配位键相互作用质谱研究新进展

顺铂及其类似物广泛应用于肿瘤的临床治疗,静脉注射顺铂后,药物通过血液传输进入细胞并最终到达细胞核作用于DNA,这一过程中,大部分的铂与血液及细胞中的蛋白质通过配位键结合,对药物的运输、代谢、累积分布和毒副作用及其生物利用度等起着决定性作用。因此,研究铂类药物与蛋白质的结合行为具有重要意义。质谱作为一种高效的蛋白质分析技术,具有灵敏度高、通量大、耗样量小、精度高等优势,在铂类药物与蛋白质相互作用研究领域发挥着重要作用。本文从Pt(Ⅱ)配合物的配位化学原理出发,阐明铂药发挥抗癌作用的可能机制。同时,着重总结、评述不同类型质谱分析方法探索铂-蛋白质配位相互作用的最新进展,归纳铂类药物在体内可能通过配位键结合的氨基酸侧链和对应的蛋白质。以顺铂为代表的Pt(Ⅱ)金属抗癌药物,最容易结合含有S供体的半胱氨酸、甲硫氨酸和含有咪唑基团的组氨酸等氨基酸侧链。同时,酪氨酸、丝氨酸、苏氨酸、谷氨酸和天冬氨酸等含羟基或羧基侧链的氨基酸也能与Pt(Ⅱ)结合。由于巯基或咪唑侧链可能处于很多蛋白质的重要活性位点或金属结合域,因此,铂类化合物的结合可能直接导致这些蛋白质的活性下降,从而对细胞内的生物过程产生复杂而深远的影响,加强或降低铂类抗癌药物的活性,或产生毒副作用和耐药性。因此,深入研究铂类药物与蛋白质的相互作用,将进一步加深对铂类抗癌药物代谢途径和毒性机制的认识,有助于设计合成更低毒、高效的金属类抗癌药物。     

LC/MS研究有机金属钌抗肿瘤化合物与蛋白酪氨酸磷酸酶模型化合物的相互作用

Protein tyrosine phosphatase 1B(PTP 1B) which contains a catalytic cysteine plays important role in the negative regulating of insulin signaling, and it has been investigated as a therapeutic target in type II diabetes and obesity. In the present work, the reactions of ruthenium anticancer complex[(ŋ⁶-cymene)Ru(en)Cl]PF₆ with the model compound(2-mercaptobenzanilide) were studied under physiologically relevant conditions. Then we treated the mono- and di-ruthenium product with GSH and H₂O₂ to mimic the inactivation and activation of PTP1B whilst [(ŋ⁶-cym)Ru(en)Cl]PF₆ binds to the active site of PTP1B. HPLC-ESI-MS time courses suggest that organometallic ruthenium complexes may inhibit the enzymatic activity of PTP1B by coordinating to the thiol in the active site, which may have important biological and pharmacologic significances in the treatment of diabetes and obesity.     

质谱技术在微塑料分析领域的发展及应用

微塑料一般指微纳米尺度的高分子聚合物，由于其普遍存在于环境中，易被生物摄入，且可能存在化学毒性，使得微塑料污染成为重要的全球环境问题。近年来，微纳米尺度塑料表征分析方法发展迅速，如粒度测定、化学鉴定和定量分析等，质谱技术已越来越多地用于微塑料的检测。本文主要介绍质谱技术在微塑料分析中的最新应用进展，阐述各类质谱及前处理技术用于微塑料定性、定量分析的原理和特点。随着质谱技术的不断发展，与其他表征手段联用方法的不断开发和应用，质谱技术将在微塑料检测领域发挥越来越重要的作用。