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| Design, Preparation, Characterization and Preliminary Evaluation of an Albumin Binding Peptide-Doxorubicin Conjugate |
| LIU Li-ping1, ZHANG Chun2, YIN Shuang2, WANG Qi2, ZHANG Yao2, YU Rong1, LIU Yong-dong2, SU Zhi-guo2 |
1 Key Laboratory of Drug Targeting and Drug Delivery System, Ministry of Education, West China School of Pharmacy, Sichuan University, Chengdu 610041, China;
2 State Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing 100190, China |
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Abstract Doxorubicin is a widely used anthracycline that has been proven highly effective in the treatment of many types of hematologic and solid cancers. The clinical application of this antitumor drug is, however, limited by its severe side effects, including neutropenia, mucositis, myelosuppression and cumulative cardiotoxicity. Besides, the therapeutic potential of this agent is also significantly limited due to the defects of short circulation time and quick clearance. In order to improve the therapeutic efficacy of doxorubicin, there is a need for developing effective strategies to prolong its blood circulation time.Currently, a novel long-term approach based on albumin is widely studied. Albumin is the most abundant and stable protein in plasma, which can maintain an extraordinarily long circulatory half-life of 15~19 days due to its size which is bigger than the renal filtration threshold and its interaction with the FcRn-mediated recycling. Based on these characteristics, albumin is thus regarded as a perfect vehicle to be used to extend the circulatory half-life of drugs which can be conjugated or genetically fused to albumin. Besides, proteins that specifically bind to albumin have also been employed to extend the half-life of therapeutic agents. Albumin-binding domain, a novel short peptide derived from bacterial surface proteins, is a naturally occurring left-handed three-helix bundle which has an extremely high affinity for HSA in the femtomolar range. It is reported that the helices 2 and 3 in surface exposed side chains of this peptide are able to selectively bind to the domainⅡ of serum albumin. The pharmacokinetic properties of many sorts of drugs can be easily improved by conjugating to this albumin-binding module. In order to improve the circulating half-life of doxorubicin in blood, a new kind of cysteine-containing albumin-binding domain was firstly expressed in E.coli and successfully prepared through a two-step chromatography with a purity of above 95%. And it is confirmed that this recombinant polypeptide had the ability of rapidly combining with human serum albumin in vitro. DOXO-EMCH, a (6-maleimidocaproyl) hydrazone derivative of doxorubicin with the thiol-sensitive maleimide group, was then conjugated to this polypeptide in a semi-organic solution through a Michael addition mechanism. The conjugation conditions including types and concentrations of organic reagent were systematically optimized and the structure characteristics of the conjugate were then preliminarily evaluated. By using A549 cells and nude mice bearing A549 xenografts as the intro and in vivo model system, the antitumor efficacy of this conjugate was tested. Furthermore, the pharmacokinetic properties in Sprague-Dawley mice were also accessed and compared with free DOX as well as DOXO-EMCH. As a result, the conjugate could be successfully synthetized with a modification yield of 73.59% when the concentration of organic phase was strictly controlled at 40%. The A549 cell cytotoxicity of the conjugate was presented with an increased IC50 value. And the conjugate exhibited a 5.6 times longer half-time than DOX, and 3.8 folds than DOXO-EMCH as was tested in the pharmacokinetic study. Besides, the growth inhibitory on tumor in vivo of this conjugate was also much stronger than those of free DOX and DOXO-EMCH at the equivalent dose. All the data indicated that the albumin-binding domain used was a promising candidate for tumor half-life extension strategy of various small antitumor drugs.
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Received: 04 November 2016
Published: 25 April 2017
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Cite this article:
LIU Li-ping, ZHANG Chun, YIN Shuang, WANG Qi, ZHANG Yao, YU Rong, LIU Yong-dong, SU Zhi-guo. Design, Preparation, Characterization and Preliminary Evaluation of an Albumin Binding Peptide-Doxorubicin Conjugate. China Biotechnology, 2017, 37(4): 68-75.
URL:
https://manu60.magtech.com.cn/biotech/10.13523/j.cb.20170409 OR https://manu60.magtech.com.cn/biotech/Y2017/V37/I4/68
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