青蒿素及其转铁蛋白复合物靶向治疗肿瘤研究进展

［摘要］研究显示，青蒿素类药物具有良好的抗肿瘤活性。由于肿瘤细胞铁含量较正常细胞高，而青蒿素可与细胞内亚铁离子产生氧化应激作用，生成大量自由基，对肿瘤细胞产生选择性杀伤作用。在青蒿素抗肿瘤作用中，转铁蛋白和转铁蛋白受体发挥重要作用。近年来，将青蒿素类药物与转铁蛋白或转铁蛋白受体结合后，可大大提高其靶向性和抗肿瘤作用，具有良好的应用前景。     

青蒿素及其衍生物的抗肿瘤作用及其最新进展

青蒿素类药物是治疗疟疾的主要药物,其衍生物有青蒿琥酯、蒿甲醚和二氢青蒿素等,主要的作用机制是通过铁离子介导的细胞损伤。近年来研究发现,青蒿素类药物还具有更广泛的药理作用,它可以通过阻滞细胞周期、诱导细胞凋亡、抗血管生成、调节肿瘤相关基因的表达以及损伤细胞线粒体等机制从而发挥抗肿瘤作用,其抗肿瘤作用日愈受到人们的重视。     

双氢青蒿素抗肿瘤作用机制的研究进展

双氢青蒿素(dihydroartemisinin,DHA)是我国自主研发的青蒿素衍生物类抗疟药,它除了具有良好的抗疟作用外,近几年研究发现其在体内外还具有较强的抗肿瘤作用。现代研究发现,DHA可以作用于线粒体依赖性细胞凋亡通路、抑制NF-κB活化,从而促进肿瘤细胞凋亡;并且能阻滞细胞周期;由Fe2+介导直接杀伤肿瘤细胞;通过作用于纤维蛋白溶解系统uPA、抑制VEGF诱导的血管生成作用来抑制肿瘤的侵袭和转移。本文对DHA抗肿瘤作用及其机制方面的研究作一综述,以期对青蒿素类药物抗癌作用研究热点提供有价值的参考。     

青蒿素类药物抗肿瘤作用的基础与临床研究

青蒿素类(Art)抗疟药具有多种药理活性,近年来对其抗肿瘤作用的基础研究较多,相关的临床研究也逐渐开展。大量体外和动物体内实验结果显示:Art可抑制或杀伤肿瘤细胞;诱导肿瘤细胞凋亡:阻滞细胞周期;抑制血管生成;延缓或逆转肿瘤细胞的多药耐药性;与铁制剂合用或与转铁蛋白结合可提高对肿增细胞的选择性杀伤作用。临床探索提示Art对肿瘤具有治疗或辅助治疗作用。Art对人体毒性低,与传统化学治疗药物有协同增效作用且无交叉耐药性。应加强Art抗肿瘤的临床研究以明确其抗肿瘤性质、范围、剂量、疗程及不良反应。     

二氢青蒿素抗肿瘤作用及其机制研究进展

二氢青蒿素是青蒿素的重要衍生物,具有显著的抗疟作用。对多种肿瘤动物模型具有一定的肿瘤抑制作用,提示其具有抗肿瘤作用。二氢青蒿素的抗肿瘤作用机制可能与抑制肿瘤细胞增殖包括诱导肿瘤细胞周期阻滞和促进肿瘤细胞凋亡、抑制肿瘤新生血管生成以及抗肿瘤细胞侵袭和转移等有关。细胞内亚铁或亚铁血红素可能是二氢青蒿素抗肿瘤作用的直接靶点。由于二氢青蒿素具有广谱抗肿瘤作用、对正常细胞毒性小及对多药耐药细胞有效等优点,因此有可能被开发为新的抗肿瘤药物。     

青蒿素及其衍生物的抗肿瘤作用

青蒿素及其衍生物是一类高效、低毒的抗疟药物 ,尤其对耐氯喹或对多种药物耐药的脑型疟和恶性疟有效。因此在临床上得到广泛的应用。随着研究的深入 ,人们发现青蒿素类化合物还具有其它很多重要的药理活性 ,如抗血吸虫、免疫调节、抗心律失常、抗肿瘤等。尤其是其抗肿瘤作用 ,越来越引起研究者们的重视。很多实验研究表明 ,青蒿素类化合物对多种肿瘤细胞的生长具有显著的抑制作用 ,而且对正常组织细胞的毒性很低 ,这为研究开发经济、有效的抗癌药物提供了新的线索。本文就近年来有关青蒿素类药物抗肿瘤的研究进展作一综述。1　青蒿素及其衍…     

青蒿素类化合物抗肿瘤机制研究- 青蒿素类化合物/转铁蛋白对接研究

本文采用柔性分子对接技术, 将11个青蒿素类化合物对接到在不同分离度下测出转铁蛋白结构的活性腔内, 研究其抗肿瘤机制, 运用多种打分函数对结果进行打分。从对接结果可看出, 转铁蛋白结构中键合铁的Asp-63、Tyr-188和His-249残基以及稳定键合位点的Arg-124和Lys-296残基与青蒿素小分子的距离小于0.5 nm, 活性大的化合物得分较高。对接后的模型解释了转铁蛋白的存在促使Fe²⁺与青蒿素作用、青蒿素不参与其他的代谢、增加青蒿素细胞毒性的机制, 为设计、合成全新青蒿素类化合物打下了良好的基础。     

受体介导肿瘤主动靶向的研究进展

肿瘤细胞表面过度表达一系列受体,能与特异性的配体或抗体结合并诱导细胞内化。以这些受体作为靶点,使药物与特异性配体或抗体结合即可将药物主动靶向肿瘤细胞。本文综述目前在临床和实验研究中用于肿瘤主动靶向的几种受体,包括酪氨酸激酶受体、叶酸受体、激素受体、脂蛋白受体、转铁蛋白受体等。通过了解这些受体及其在肿瘤主动靶向方面的作用,有助于我们对抗肿瘤药物和新型抗肿瘤药物载体的研究。     

受体介导肿瘤主动靶向的研究进展

肿瘤细胞表面过度表达一系列受体，能与特异性的配体或抗体结合并诱导细胞内化。以这些受体作为靶点，使药物与特异性配体或抗体结合即可将药物主动靶向肿瘤细胞。本文综述目前在临床和实验研究中用于肿瘤主动靶向的几种受体，包括酪氨酸激酶受体、叶酸受体、激素受体、脂蛋白受体、转铁蛋白受体等。通过了解这些受体及其在肿瘤主动靶向方面的作用，有助于我们对抗肿瘤药物和新型抗肿瘤药物载体的研究。     

受体介导的肿瘤靶向治疗研究进展

肿瘤细胞表面过度表达一系列受体，能与特异性的配体结合并诱导细胞内化。以这些受体为作用靶点，使抗肿瘤药物与特异性配体结合即可将药物主动靶向肿瘤细胞。本文就近年来研究较多的受体如唾液酸糖蛋白受体、生长因子受体、低密度酯蛋白受体、转铁蛋白受体、叶酸受体、CD44等进行概述。     

Anticancer properties of artemisinin derivatives and their targeted delivery by transferrin conjugation

Artemisinin and its derivatives are well known antimalaria drugs and particularly useful for the treatment of infection of Plasmodium falciparum malaria parasites resistant to traditional antimalarials. Artemisinin has an endoperoxide bridge that is activated by intraparasitic heme-iron to form free radicals, which kill malaria parasites by alkylating biomolecules. In recent years, there are many reports of anticancer activities of artemisinins both in vitro and in vivo. Artemisinins have inhibitory effects on cancer cell growth, including many drug- and radiation-resistant cancer cell lines. The cytotoxic effect of artemisinin is specific to cancer cells because most cancer cells express a high concentration of transferrin receptors on cell surface and have higher iron ion influx than normal cells via transferrin mechanism. In addition, some artemisinin analogs have been shown to have antiangiogenesis activity. Artemisinin tagged to transferrin via carbohydrate chain has also been shown to have high potency and specificity against cancer cells. The conjugation enables targeted delivery of artemisinin into cancer cells. In this review, we discuss the anticancer activities and mechanisms of action of artemisinins and the transferrin-conjugate.     

Targeted treatment of cancer with artemisinin and artemisinin-tagged iron-carrying compounds

Artemisinin is a chemical compound that reacts with iron to form free radicals which can kill cells. Cancer cells require and uptake a large amount of iron to proliferate. They are more susceptible to the cytotoxic effect of artemisinin than normal cells. Cancer cells express a large concentration of cell surface transferrin receptors that facilitate uptake of the plasma iron-carrying protein transferrin via endocytosis. By covalently tagging artemisinin to transferrin, artemisinin could be selectively picked up and concentrated by cancer cells. Futhermore, both artemisinin and iron would be transported into the cell in one package. Once an artemisinin-tagged transferrin molecule is endocytosed, iron is released and reacts with artemisinin moieties tagged to transferrin. Formation of free radicals kills the cancer cell. The authors have found that artemisinin-tagged transferrin is highly selective and potent in killing cancer cells. Thus, artemisinin and artemisinin-tagged iron-carrying compounds could be developed into powerful anticancer drugs.     

Targeted treatment of cancer with artemisinin and artemisinin-tagged iron-carrying compounds

Artemisinin is a chemical compound that reacts with iron to form free radicals which can kill cells. Cancer cells require and uptake a large amount of iron to proliferate. They are more susceptible to the cytotoxic effect of artemisinin than normal cells. Cancer cells express a large concentration of cell surface transferrin receptors that facilitate uptake of the plasma iron-carrying protein transferrin via endocytosis. By covalently tagging artemisinin to transferrin, artemisinin could be selectively picked up and concentrated by cancer cells. Futhermore, both artemisinin and iron would be transported into the cell in one package. Once an artemisinin-tagged transferrin molecule is endocytosed, iron is released and reacts with artemisinin moieties tagged to transferrin. Formation of free radicals kills the cancer cell. The authors have found that artemisinin-tagged transferrin is highly selective and potent in killing cancer cells. Thus, artemisinin and artemisinin-tagged iron-carrying compounds could be developed into powerful anticancer drugs.     

Oxidative stress response of tumor cells: microarray-based comparison between artemisinins and anthracyclines

The antimalarial artemisinins also reveal profound cytotoxic activity against tumor cells. Artemisinins harbor an endoperoxide bridge whose cleavage results in the generation of reactive oxygen species (ROS) and/or artemisinin carbon-centered free radicals. Established cancer drugs such as anthracyclines also form ROS and free radicals that are responsible for the cardiotoxicity of anthracyclines. In contrast, artemisinins do not reveal cardiotoxicity. In the present investigation, we compared the cytotoxic activities of different artemisinins (artemisinin, artesunate, arteether, artemether, artemisitene, dihydroartemisinylester stereoisomers) in 60 cell lines of the National Cancer Institute (NCI), USA, with those of anthracyclines (doxorubicin, daunorubicin, 4'-epirubicin, idarubicin, deoxydoxorubicin, trifluoroacetyl-doxorubicin-14-valerate). The inhibition concentration 50% (IC(50)) values of artemisinins and anthracyclines were correlated with the mRNA expression of 170 genes involved in oxygen stress response and metabolism as recently determined by microarray analysis and deposited in the NCI database (http://dtp.nci.nih.gov). The genes whose expression was significantly linked to cellular drug response in Kendall's tau tests were subjected to hierarchical cluster analysis and cluster image mapping. Mathematical correction for false-positive correlations was done by a false discovery rate algorithm. One cluster contained predominantly genes with a relationship to artemisinins and another one genes with a relationship to anthracyclines. In a third cluster, genes correlating to both drug classes were assembled. This indicates that different sets of genes involved in oxidative stress response and metabolism may contribute to the cytotoxic and differing toxic side effects of these drug classes.     

Molecular pharmacology and pharmacogenomics of artemisinin and its derivatives in cancer cells

Secondary metabolites from plants can serve as defense against herbivores, microbes, viruses or competing plants. Many compounds from medicinal plants have pharmacological activities and thus may be a source for novel anti-tumor agents. We have analyzed natural products from traditional Chinese medicine during the past decade and focused our interest on the compound artemisinin from Artemisia annua L. (qinghao, sweet wormwood) and its derivatives. In addition to their anti-malarial properties, artemisinins are cytotoxic for cancer cells. The present review focuses on the mechanisms of action of artemisinins in cancer cells relating to: 1. anti-proliferative and anti-angiogenic effects, 2. induction of apoptosis, 3. oxidative stress, 4. oncogenes and tumor suppressor genes, and 5. multidrug resistance. Data on putative target molecules of artemisinins are presented and discussed, e.g. the translationally controlled tumor protein (TCTP). Emphasis is given to pharmacogenomic approaches to analyze the pleiotropic nature of mechanisms of artemisinins in cancer cells.     

Dihydroartemisinin inhibits the human erythroid cell differentiation by altering the cell cycle

Artemisinin derivatives such as dihydroartemisinin (DHA) induce significant depletion of early embryonic erythroblasts in animal models. We have reported previously that DHA specifically targets pro-erythroblasts and basophilic erythroblasts, when human CD34+ stem cells are differentiated toward the erythroid lineage, indicating that a window of susceptibility to artemisinins may exist also in human developmental erythropoiesis during pregnancy. To better investigate the toxicity of artemisinin derivatives, the structure-activity relationship was evaluated against the K562 leukaemia cell line, used as a model for differentiating early human erythroblasts. All artemisinins derivatives, except deoxyartemisinin, inhibited both spontaneous and induced erythroid differentiation, confirming that the peroxide bridge is responsible for the erythro-toxicity. On the contrary, cell growth was markedly reduced by DHA, artemisone and artesunate but not by artemisinin, 10-deoxoartemisinin or deoxy-artemisinin. The substituent at position C-10 is responsible only for the anti-proliferative effect, since 10-deoxoartemisinin did not reduce cell growth but arrested the differentiation of K562 cells. In particular, the results showed that DHA resulted the most potent and rapidly acting compound of the drug family, causing (i) the decreased expression of GpA surface receptors and the down regulation the γ-globin gene; (ii) the alteration of S phase of cell cycle and (iii) the induction of programmed cell death of early erythroblasts in a dose dependent manner within 24h. In conclusion, these findings confirm that the active metabolite DHA is responsible for the erythro-toxicity of most of artemisinins used in therapy. Thus, as long as no further clinical data are available, current WHO recommendations of avoiding malaria treatment with artemisinins during the first trimester of pregnancy remain valid.     

Artemisinin Antimalarials: Preserving the "Magic Bullet"

The artemisinins are the most effective antimalarial drugs known. They possess a remarkably wide therapeutic index. These agents have been used in traditional Chinese herbal medicine for more than 2,000 years but were not subjected to scientific scrutiny until the 1970s. The first formal clinical trials of the artemisinins, and the development of methods for their industrial scale production, followed rapidly. A decade later, Chinese scientists shared their findings with the rest of the world; since then, a significant body of international trial evidence has confirmed these drugs to be far superior to any available alternatives. In particular, they have the ability to rapidly kill a broad range of asexual parasite stages at safe concentrations that are consistently achievable via standard dosing regimens. As their half-life is very short, there was also thought to be a low risk of resistance. These discoveries coincided with the appearance and spread of resistance to all the other major classes of antimalarials. As a result, the artemisinins now form an essential element of recommended first-line antimalarial treatment regimens worldwide. To minimize the risk of artemisinin resistance, they are recommended to be used to treat uncomplicated malaria in combination with other antimalarials as artemisinin combination therapies (ACTs). Their rollout has resulted in documented reductions in malaria prevalence in a number of African and Asian countries. Unfortunately, there are already worrisome early signs of artemisinin resistance appearing in western Cambodia. If this resistance were to spread, it would be disastrous for malaria control efforts worldwide. The enormous challenge for the international community is how to avert this catastrophe and preserve the effectiveness of this antimalarial "magic bullet". Drug Dev Res 71: 12-19, 2010. ? 2009 Wiley-Liss, Inc.     

Preparation,characterization and in vitro efficacy of magnetic nanoliposomes containing the artemisinin and transferrin

Background

Artemisinin is the major sesquiterpene lactones in sweet wormwood (Artemisia annua L.), and its combination with transferrin exhibits versatile anti-cancer activities. Their non-selective targeting for cancer cells, however, limits their application. The aim of this study was to prepare the artemisinin and transferrin-loaded magnetic nanoliposomes in thermosensitive and non-thermosensitive forms and evaluate their antiproliferative activity against MCF-7 and MDA-MB-231 cells for better tumor-targeted therapy.

Methods

Artemisinin and transferrin-loaded magnetic nanoliposomes was prepared by extrusion method using various concentrations of lipids. These formulations were characterized for particle size, zeta potential, polydispersity index and shape morphology. The artemisinin and transferrin-loading efficiencies were determined using HPLC. The content of magnetic iron oxide in the nanoliposomes was analysed by spectrophotometry. The in vitro release of artemisinin, transferrin and magnetic iron oxide from vesicles was assessed by keeping of the nanoliposomes at 37°C for 12 h. The in vitro cytotoxicity of prepared nanoliposomes was investigated against MCF-7 and MDA-MB-231 cells using MTT assay.

Results

The entrapment efficiencies of artemisinin, transferrin and magnetic iron oxide in the non-thermosensitive nanoliposomes were 89.11% ± 0.23, 85.09% ± 0.31 and 78.10% ± 0.24, respectively. Moreover, the thermosensitive formulation showed a suitable condition for thermal drug release at 42°C and exhibited high antiproliferative activity against MCF-7 and MDA-MB-231 cells in the presence of a magnetic field.

Conclusions

Our results showed that the thermosensitive artemisinin and transferrin-loaded magnetic nanoliposomes would be an effective choice for tumor-targeted therapy, due to its suitable stability and high effectiveness.     

Are currently deployed artemisinins neurotoxic?

In vitro, animal, and human clinical studies suggest currently deployed artemisinins possess neurotoxic potential. A specific and consistent pattern of brainstem injuries that includes auditory processing centers has been reported from all laboratory animals studied. Hearing loss, ataxia, and tremor are reported from humans. Neurotoxicity appears mediated in part through artemisinin induced oxidative stress in exposed brainstems. In vitro studies suggest that artemisinin neurotoxicity does not manifest immediately upon exposure, but that once commenced it is inevitable and irreversible; extrapolation from in vitro data suggests that 14 days may possibly be required for full development, casting doubt upon some animal safety studies and human necropsy studies. Uncertainty remains over the neurotoxicity of currently deployed artemisinins, and their safety profile should be reviewed, especially in pediatric use. The development of non-neurotoxic artemisinins is possible and should be encouraged.