抗肿瘤多肽类药物的来源及其机制的研究进展

恶性肿瘤已成为严重影响人类健康的一种疾病,其治疗手段一直是人们所探索的内容,药物治疗就是其中一种治疗方法。与传统化疗药物相比,抗肿瘤多肽类药物以其分子量小、特异性强、毒性低等特点作为新的治疗肿瘤药物一直广受人们关注。多肽的来源广泛,有存在于天然动植物及微生物体内,也可以通过蛋白质酶解或人工合成得到。近年来,越来越多的多肽类药物被发现除抗菌外还具有抗肿瘤的作用,其抗肿瘤的机制多种多样但尚未完全清楚,是许多研究者研究的重点。本文从多肽类药物的来源与特点及它们的抗肿瘤机制等方面对多肽类药物的研究进展进行综述。     

胃旁路术对“肠-胰岛轴”相关激素及二肽基肽酶-IV分泌的影响

目的:探讨胃旁路术(GBP)在降低2型糖尿病(T2DM)大鼠血糖过程中对“肠-胰岛轴”相关激素及二肽基肽酶Ⅳ( DPP-Ⅳ)分泌的影响.方法:将24只雄性自发性非肥胖型糖尿病(GK)大鼠随机均分为胃旁路组和假手术组,记录并对比两组大鼠术前及术后1,2,4,8,12,24,48周体质量、空腹血糖及血浆DPP-Ⅳ、“肠-胰岛轴”相关激素胰高血糖素样肽1( GLP-1)水平.结果:术前两组大鼠间各项观察指标无明显差异(均P＞0.05);两组大鼠体质量均在术后1周内明显降低,但随后逐渐回升,且两组间体质量在术后各时间点均无统计学差异(均P＞0.05);与术前比较,假手术组大鼠术后空腹血糖及血浆DPP-Ⅳ,GLP-1水平无明显变化,胃旁路组空腹血糖及血浆DPP-Ⅳ水平明显降低,而GLP-1水平明显增高(均P＜0.01),且两组间空腹血糖及血浆DPP-Ⅳ,GLP-1水平在术后各时间点上的差异均有统计学意义(均P＜0.01).结论:胃旁路术控制血糖的作用可能与抑制DPP-Ⅳ的活性,使机体中“肠-胰岛轴”上GLP-1水平升高有关.     

3种人血清白蛋白降解多肽片段的血液相容性研究

目的 研究3种人血清白蛋白降解多肽片段(PF1-123、PF124-298、PF299-585)的血液相容性.方法 分别考察3种多肽片段对新鲜人血的溶血率、对血细胞形态的影响、全血凝固时间以及血浆复钙时间,测试其血液相容性.结果 PF1-123、PF124-298和PF299-585对新鲜人血的溶血率分别为0.52％±0.47％、0.39％±0.33％、0.32％±0.23％,对血细胞形态均无不良影响,说明3种多肽片段均符合医用材料的溶血率要求;3种多肽片段的全血凝固时间曲线以及血浆复钙时间曲线与人血清白蛋白相似,说明其与血液接触后不会激活相关因子而促成凝血.结论 PF1-123、PF124-298和PF299-585具有良好的血液相容性.     

Nanoparticle delivery of CDDO-Me remodels the tumor microenvironment and enhances vaccine therapy for melanoma

Lipid-calcium-phosphate nanoparticle (NP) delivery of Trp2 peptide vaccine is one of the most effective vaccine strategies against melanoma. However, due to the immunosuppressive microenvironment in the tumor, the achievement of potent immune responses remains a major challenge. NP delivery systems provide an opportunity to deliver chemotherapy agent to modulate the tumor microenvironment (TME) and improve the vaccine activity. Anti-inflammatory triterpenoid methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) is a broad spectrum inhibitor of several signaling pathways that are important in both cancer cells and cells in the TME. Intravenous delivery of CDDO-Me using poly-lactic-glycolic-acid NP combination with subcutaneous Trp2 vaccine resulted in an increase of antitumor efficacy and apoptotic tumor tissue than Trp2 vaccine alone in B16F10 melanoma. There was a significant decrease of both Treg cells and MDSCs and a concomitant increase in the cytotoxic T-lymphocyte infiltration in TEM of the vaccinated animals. Also, CDDO-Me remodeled the tumor associated fibroblasts, collagen and vessel in TME, meanwhile, enhanced the Fas signaling pathway which could sensitize the tumor cells for cytotoxic T lymphocyte mediated killing. The combination of systemic induction of antigen-specific immune response using Trp2 nanovaccine and targeted modification of the TME with the NP delivered CDDO-Me offers a powerful combination therapy for melanoma.     

Proteomic Analysis for Finding Serum Pathogenic Factors and Potential Biomarkers in Multiple Myeloma

Background:

Multiple myeloma (MM) is a malignant tumor, which takes the second place in malignant blood disease. The clinical symptoms are complicated that make more difficult to diagnose and therapy. Lots of researches focus on the proteins about MM in order to solve those problems. We used proteomic methods to find potential biomarkers in MM patients.

Methods:

We applied the peptide ligand library beads (PLLBs) to deplete high abundance proteins in serum for finding potential pathogenic factors and biomarkers of MM. Using 1D-Gel-liquid chromatography-tandem mass spectrometry (LC-MS/MS), we identified 789 and 849 unique serum proteins in MM patients and in healthy controls, respectively.

Results:

Twenty-two proteins were found differentially expressed between the two groups including serum amyloid A protein, vitamin D-binding protein isoform-1 precursor, plasma kallikrein, and apolipoprotein A-I. Changes of integrin alpha-11 and isoform-1 of multimerin-1 were validated with Western blotting. The linkage of the differentially expressed proteins and the pathogenesis pathways of MM were discussed.

Conclusions:

PLLB combined with 1D-gel-LC-MS/MS analysis is an efficient method to identify differentially expressed proteins in serum from patients with MM.     

Localized delivery of mechano-growth factor E-domain peptide via polymeric microstructures improves cardiac function following myocardial infarction

The Insulin like growth factor-I isoform mechano-growth factor (MGF), is expressed in the heart following myocardial infarction and encodes a unique E-domain region. To examine E-domain function, we delivered a synthetic peptide corresponding to the unique E-domain region of the human MGF (IGF-1Ec) via peptide eluting polymeric microstructures to the heart. The microstructures were made of poly (ethylene glycol) dimethacrylate hydrogel and bioengineered to be the same size as an adult cardiac myocyte (100 × 15 × 15 μm) and with a stiffness of 20 kPa. Peptide eluting microrods and empty microrods were delivered via intramuscular injection following coronary artery ligation in mice. To examine the physiologic consequences, we assessed the impact of peptide delivery on cardiac function and cardiovascular hemodynamics using pressure–volume loops and gene expression by quantitative RT-PCR. A significant decline in both systolic and diastolic function accompanied by pathologic hypertrophy occurred by 2 weeks which decompensated further by 10 weeks post-infarct in the untreated groups. Delivery of the E-domain peptide eluting microrods decreased mortality, ameliorated the decline in hemodynamics, and delayed decompensation. This was associated with the inhibition of pathologic hypertrophy despite increasing vascular impedance. Delivery of the empty microrods had limited effects on hemodynamics and while pathologic hypertrophy persisted there was a decrease in ventricular stiffness. Our data show that cardiac restricted administration of the MGF E-domain peptide using polymeric microstructures may be used to prevent adverse remodeling of the heart and improve function following myocardial infarction.     

Design of self-assembling peptide hydrogelators amenable to bacterial expression

Hydrogels formed from self-assembling peptides are finding use in tissue engineering and drug delivery applications. Given the notorious difficulties associated with producing self-assembling peptides by recombinant expression, most are typically prepared by chemical synthesis. Herein, we report the design of a family of self-assembling β-hairpin peptides amenable to efficient production using an optimized bacterial expression system. Expressing peptides, EX1, EX2 and EX3 contain identical eight-residue amphiphilic β-strands connected by varying turn sequences that are responsible for ensuring chain reversal and the proper intramolecular folding and consequent self-assembly of the peptide into a hydrogel network under physiological conditions. EX1 was initially used to establish and optimize the bacterial expression system by which all the peptides could be eventually individually expressed. Expression clones were designed to allow exploration of possible fusion partners and investigate both enzymatic and chemical cleavage as means to liberate the target peptide. A systematic analysis of possible expression systems followed by fermentation optimization lead to a system in which all three peptides could be expressed as fusions with BAD-BH3, the BH3 domain of the proapoptotic BAD (Bcl-2 Associated Death) Protein. CNBr cleavage followed by purification afforded 50, 31, and 15 mg/L yields of pure EX1, EX2 and EX3, respectively. CD spectroscopy, TEM, and rheological analysis indicate that these peptides fold and assembled into well-defined fibrils that constitute hydrogels having shear-thin/recovery properties.     

Relative contribution of different l-arginine sources to the substrate supply of endothelial nitric oxide synthase

In certain cases of endothelial dysfunction l-arginine becomes rate-limiting for NO synthesis in spite of sufficiently high plasma concentrations of the amino acid. To better understand this phenomenon, we investigated routes of substrate supply to endothelial nitric oxide synthase (eNOS). Our previous data with human umbilical vein (HUVEC) and EA.hy.926 endothelial cells demonstrated that eNOS can obtain its substrate from the conversion of l-citrulline to l-arginine and from protein breakdown. In the present study, we determined the quantitative contribution of proteasomal and lysosomal protein degradation and investigated to what extent extracellular peptides and l-citrulline can provide substrate to eNOS. The RFL-6 reporter cell assay was used to measure eNOS activity in human EA.hy926 endothelial cells. Individual proteasome and lysosome inhibition reduced eNOS activity in EA.hy926 cells only slightly. However, the combined inhibition had a pronounced reducing effect. eNOS activity was fully restored by supplementing either l-citrulline or l-arginine-containing dipeptides. Histidine prevented the restoration of eNOS activity by the dipeptide, suggesting that a transporter accepting both, peptides and histidine, mediates the uptake of the extracellular peptide. In fact, the peptide and histidine transporter PHT1 was expressed in EA.hy926 cells and HUVECs (qRT/PCR). Our study thus demonstrates that l-citrulline and l-arginine-containing peptides derived from either intracellular protein breakdown or from the extracellular space seem to be good substrate sources for eNOS.     

Variants of self-assembling peptide,KLD-12 that show both rapid fracture healing and antimicrobial properties

KLD-12 (KLD) is a 12-residue self-assembling peptide that can adopt nano-structures and is known for its tissue-engineering properties. Our objective was to introduce antimicrobial attribute to KLD which would help in preventing secondary infection associated with external application of such tissue engineering materials. Considering the net charge of KLD-12, varying number of cationic arginine residues were added to its N-terminus. KLD variants showed appreciable bactericidal properties without any significant increase in cytotoxicity against tested mammalian cells. Further, these variants adopted β-sheet structures and self-assembled into nano-structures comparable to that of KLD. Interestingly, the KLD variants with two (KLD-2R) and three (KLD-3R) arginine residues added to its N-terminus showed significant osteogenic effect which was comparable or better than the original peptide as evident from the alkaline phosphatase activity assay, mineralized nodule formation and expression of different osteogenic genes. Particularly, application of KLD-2R in rats to the site of a drill-hole (0.8 mm diameter) that was created in the femur metaphysis displayed significantly higher bone regeneration compared to that of KLD. The results demonstrate a simple way to improve biological property of a self-assembling peptide with tissue engineering property.     

奥瑞凝胶对反流性食管炎大鼠模型食管组织中相关基因表达及血清相关分子含量的影响

目的:研究奥瑞凝胶对反流性食管炎大鼠模型食管组织中相关基因表达以及血清中相关分子含量的影响.方法:选择成年雄性SD大鼠作为研究对象,随机分为正常组、模型组和治疗组,模型组和治疗组建立反流性食管炎模型,治疗组给予奥瑞凝胶治疗.处死大鼠后,检测血清炎症因子含量和食管组织炎症因子、多肽类神经递质、促增殖基因的表达.结果:(1)炎症因子:与模型组比较,食管组织和血清中白介素-23、17(IL-23、IL-17)的含量在治疗组中呈降低趋势;(2)多肽类神经递质:与模型组比较,食管组织中一氧化氮(NO)、NOS、血管活性肠肽(VIP)、VIP-R1、VIP-R2含量在治疗组中呈升高趋势,P物质(SP)含量治疗组中呈降低趋势;(3)促增殖基因:与模型组比较,食管组织中c-myb、增殖细胞核抗原(PCNA)和Ki-67的mRNA和蛋白含量在治疗组中呈降低趋势.结论:奥瑞凝胶治疗有助于缓解炎症反应,调节多肽类神经递质表达,抑制食管黏膜上皮过度增殖,对反流性食管炎模型大鼠具有治疗作用.