提高免疫应答的新佐剂与投递系统

<正>根据美国剑桥科学家的近期研究,疫苗是预防传染病的最重要的手段,在有效疫苗的研发中,除了鉴定出最相关免疫原和有效的接种方案是主要的挑战外,选择有效的佐剂和投递方法同样是关键性的问题。免疫原与不同的佐剂配方对抗体应答的亲和     

电脉冲和布吡卡因增强A型肉毒毒素DNA核酸疫苗的免疫效果

目的：考查DNA疫苗注射免疫后电脉冲和布吡卡因佐剂化DNA疫苗递送方式对A型肉毒毒素DNA核酸疫苗免疫效果的影响。方法：A型肉毒毒素DNA复制子疫苗和传统DNA疫苗肌肉注射免疫小鼠后电脉冲和布吡卡因佐剂化DNA后再肌肉注射免疫小鼠;检测免疫小鼠的抗体和细胞水平,并分析抗体亚类。结果：电脉冲和布吡卡因这二种递送方式均增强DNA复制子疫苗和传统DNA疫苗的体液免疫和细胞免疫效果;电脉冲提高DNA疫苗免疫效果更为明显,并且电脉冲和布吡卡因组合这种递送方式增强DNA疫苗体液免疫和细胞免疫水平最高;与传统DNA疫苗相比,A型肉毒毒素DNA复制子疫苗在这些递送方式下均诱导产生了更好的特异性体液免疫和细胞免疫应答,并且这些递送方式没有改变DNA疫苗的Th1/Th2免疫应答特性,即DNA复制子疫苗诱导产生Th1/Th2混合免疫应答但偏向于Th2途经,而传统DNA疫苗则完全偏向于Th2途经。结论：电脉冲和布吡卡因增强DNA复制子疫苗和传统DNA疫苗的免疫效果,是提高DNA疫苗免疫原性的良好策略。     

HEVAg-PLGA纳米颗粒疫苗小鼠体内免疫学研究

研究重组戊型肝炎抗原(HEVAg)-乳酸/乙醇酸共聚物(PLGA)纳米颗粒抗原能否在动物体内诱导产生免疫应答。制备HEVAg-PLGA纳米颗粒抗原后,通过皮下、滴鼻、口服途径接种Balb/c小鼠,每隔4周加强免疫两次,HEVAg与铝盐佐剂(铝佐剂疫苗Al_2O_3-Ag)为对照组,一定时间内检测抗体及细胞因子的应答水平。结果HEVAg-PLGA纳米颗粒抗原在小鼠体内诱导产生有效的体液免疫、细胞免疫。滴鼻、口服途径黏膜系统中诱导产生较高滴度的IgA抗体,ELISPOT结果显示鼻腔、唾液腺中IgA ASCs数量显著增加;皮下途径诱导产生较高滴度的IgG抗体;常规铝佐剂疫苗相比于HEVAg-PLGA纳米颗粒抗原诱导较强的IgG抗体水平,未诱导产生黏膜免疫应答;HEVAg-PLGA纳米颗粒抗原诱导产生较强细胞免疫应答,皮下接种途径IFN-γ、IL-4生成细胞数量显著高于其它免疫组。与铝佐剂疫苗相比,HEVAg-PLGA纳米颗粒抗原能有效诱导产生系统免疫及黏膜免疫应答,显示HEVAg-PLGA有潜力成为备选HEV黏膜疫苗抗原,同时展示PLGA颗粒作为黏膜系统抗原递送载体及黏膜佐剂的优越性。     

流感新型黏膜疫苗的研究

目的评价PorA、PorB和Class4对流感裂解疫苗的免疫增强作用,从中挑选出最有效的流感黏膜佐剂,为发展流感黏膜疫苗提供理论基础。方法流感三价裂解抗原按比例与PorA、PorB和Class4非共价结合,滴鼻免疫Balb／c小鼠3次,采取间接ELISA检测血清特异性IgG抗体及抗体亚型,检测鼻咽、肺、小肠和阴道冲洗液中IgA效价,采用血凝抑制试验检测血清中HAI效价。结果PorB重组蛋白佐剂组较无佐剂的流感裂解抗原组在提高小鼠早期免疫应答的同时诱导较强的系统免疫应答和黏膜免疫应答;PorA组也有黏膜佐剂的功能,但和无佐剂的流感裂解抗原组相比,差异无统计学意义。结论在蛋白体的三分子中,以PorB为佐剂的流感黏膜疫苗不仅提高了抗原的系统免疫应答,而且诱导了较强的小鼠呼吸道、生殖道的局部黏膜免疫应答,为流感黏膜疫苗的研制奠定了理论基础。     

纳米疫苗在肿瘤免疫疗法中的应用

近年来肿瘤免疫疗法成为癌症治疗领域的热点,其中结合肿瘤疫苗和纳米技术的纳米疫苗为肿瘤免疫疗法提供了新思路.纳米疫苗可以实现疫苗和佐剂的共载,且智能化的纳米载体进一步实现了抗原有效的靶向递送,促进了抗原的摄取和递呈,激活抗原特异性免疫应答,有效杀伤肿瘤细胞.本文就纳米疫苗的原理、优势、纳米材料的类型、临床疗效进行综述,为后期纳米疫苗的设计提供更可靠的参考依据.     

双特异抗体定向运载抗原作为一种新型佐剂系统的研究进展

目前采用基因重组等生物技术制备了大量的纯化亚单位疫苗或合成疫苗,这些疫苗的免疫原性很弱,需要佐剂的帮助才能有效地引发抗体应答。传统的佐剂具有很多本身无法克服的缺陷,促使人们研究开发新的佐剂系统。本文对双特异抗体的概念、Ｔ细胞在免疫应答中的作用及抗原的定向运送、双特异抗体介导的抗原定向运送及佐剂效应等方面对双特异抗体定向运送抗原作为一个新的佐剂系统给予了较为详细的阐述     

疫苗输送系统研究进展

随着疫苗学的发展,疫苗配方出现改进,疫苗内除含有抗原与佐剂成份外,还有输送系统.疫苗输送系统对于蛋白亚单位、核酸等疫苗的作用尤为重要,其除具有与其它佐剂一样能增强免疫应答外,可以较长时间内刺激动物免疫系统,减少免疫次数,从而降低费用.疫苗输送系统对开发防控人类和动物疫病的新型疫苗具有重要的理论与现实意义.笔者就近年来疫苗输送系统的研究进展做一综述.     

ISA 61 VG佐剂增强单核细胞增生李斯特氏菌灭活疫苗的保护性免疫应答

单核细胞增生李斯特氏菌(Listeria monocytogenes,Lm)是重要的人兽共患李斯特氏菌病的致病菌,疫苗免疫是预防该病原菌感染的有效手段之一。本研究研制了添加矿物油佐剂MontanideTM ISA61VG的新型灭活细菌疫苗,并对其安全性和免疫应答特性进行了研究。结果表明,ISA 61 VG佐剂疫苗具有较好的安全性;诱导小鼠产生的抗李斯特氏菌溶血素O抗体滴度以及IgG2a/IgG1比值显著高于无佐剂免疫组;在致死剂量Lm攻毒下,能对小鼠提供100%的免疫保护。因此,ISA 61VG佐剂能显著增强灭活疫苗诱导宿主产生体液免疫和细胞免疫应答的能力,从而提高灭活疫苗的保护性免疫应答作用,是预防人和动物Lm感染的潜在疫苗候选株。     

黏膜免疫与结核疫苗研发CSCD

结核病是全球重要的传染性疾病之一,在全球范围内保持着较高的发病率和死亡率。卡介苗是目前临床上唯一应用的结核疫苗,虽然对儿童有较好的保护作用,但对成人的免疫保护效果并不明显。研发新的结核疫苗对于结核病的防控具有重要的意义。由于结核病的致病菌结核分枝杆菌主要通过呼吸道传播,机体的黏膜成为抵御结核分枝杆菌的第一道防线。设计稳定高效的抗结核黏膜免疫疫苗是目前结核疫苗研究的新方向之一。选择合适的黏膜免疫途径、佐剂及抗原递送系统是黏膜疫苗研发成功的关键。本文对抗结核分枝杆菌的黏膜免疫应答作简短的概述,并重点阐明黏膜免疫在结核疫苗研发中的研究进展。     

新型疫苗佐剂的研究进展

与传统的灭活或活体疫苗相比,由基因工程重组抗原或化学合成多肽组成的现代疫苗往往存在免疫原性弱等问题,需要新型的免疫佐剂来增强其作用。尽管传统的铝盐佐剂是目前唯一全球公认的人用佐剂,但存在激发细胞免疫应答能力差等不足,因此,需要研发更为安全有效的人用新型佐剂,尤其是安全无毒、能够刺激较强细胞免疫应答的佐剂,以及适合粘膜疫苗、DNA疫苗和癌症疫苗的免疫佐剂。分析阐述了新型佐剂研究状况和佐剂发展方向,并进一步对新型佐剂的临床前和临床试验研究以及已批准上市的新型疫苗佐剂进行了综述。     

Lipid and carbohydrate based adjuvant/carriers in immunology.

This review discusses various issues regarding vaccines; what are they and how they work, safety aspects, the role of adjuvants and carriers in vaccination, synthetic peptides as immunogens, and new technologies for vaccine development and delivery including the identification of novel adjuvants for mucosal vaccine delivery. There has been a recent increase of interest in the use of lipids and carbohydrates as adjuvants, and so a particular emphasis is placed on adjuvants derived from lipids or carbohydrates, or from both.     

Innate endogenous adjuvants prime to desirable immune responses via mucosal routes

Vaccination is an effective strategy to prevent infectious or immune related diseases, which has made remarkable contribution in human history. Recently increasing attentions have been paid to mucosal vaccination due to its multiple advantages over conventional ways. Subunit or peptide antigens are more reasonable immunogens for mucosal vaccination than live or attenuated pathogens, however adjuvants are required to augment the immune responses. Many mucosal adjuvants have been developed to prime desirable immune responses to different etiologies. Compared with pathogen derived adjuvants, innate endogenous molecules incorporated into mucosal vaccines demonstrate prominent adjuvanticity and safety. Nowadays, cytokines are broadly used as mucosal adjuvants for participation of signal transduction of immune responses, activation of innate immunity and polarization of adaptive immunity. Desired immune responses are promptly and efficaciously primed on basis of specific interactions between cytokines and corresponding receptors. In addition, some other innate molecules are also identified as potent mucosal adjuvants. This review focuses on innate endogenous mucosal adjuvants, hoping to shed light on the development of mucosal vaccines.     

Antigenic epitopes fused to cationic peptide bound to oligonucleotides facilitate Toll-like receptor 9-dependent, but CD4+ T cell help-independent, priming of CD8+ T cells

A priority in current vaccine research is the development of adjuvants that support the efficient priming of long-lasting, CD4(+) T cell help-independent CD8(+) T cell immunity. Oligodeoxynucleotides (ODN) with immune-stimulating sequences (ISS) containing CpG motifs facilitate the priming of MHC class I-restricted CD8(+) T cell responses to proteins or peptides. We show that the adjuvant effect of ISS(+) ODN on CD8(+) T cell priming to large, recombinant Ag is enhanced by binding them to short, cationic (arginine-rich) peptides that themselves have no adjuvant activity in CD8(+) T cell priming. Fusing antigenic epitopes to cationic (8- to 10-mer) peptides bound to immune-stimulating ISS(+) ODN or nonstimulating NSS(+) ODN (without CpG-containing sequences) generated immunogens that efficiently primed long-lasting, specific CD8(+) T cell immunity of high magnitude. Different MHC class I-binding epitopes fused to short cationic peptides of different origins showed this adjuvant activity. Quantitative ODN binding to cationic peptides strikingly reduced the toxicity of the latter, suggesting that it improves the safety profile of the adjuvant. CD8(+) T cell priming supported by this adjuvant was Toll-like receptor 9 dependent, but required no CD4(+) T cell help. ODN (with or without CpG-containing sequences) are thus potent Th1-promoting adjuvants when bound to cationic peptides covalently linked to antigenic epitopes, a mode of Ag delivery prevailing in many viral nucleocapsids.     

Immunization against multiple viruses by using solid-matrix-antibody-antigen complexes.

We have previously shown that mice immunized with solid-matrix-antibody-antigen (SMAA) complexes in the absence of adjuvants show vigorous humoral and cell-mediated immune responses to the immunizing antigen. Here we report that various proteins involved in inducing protective immune responses to different viruses can easily and simply be incorporated into SMAA complexes and that such complexes act as powerful multivalent immunogens. Construction of such SMAA complexes may be one of the most practical and effective ways of producing multivalent subunit vaccines for use in humans and animals.     

Synthetic peptide vaccines

The review considers the stages of the development of synthetic peptide vaccines against infectious agents, novel approaches and technologies employed in this process, including bioinformatics, genomics, proteomics, large-scale peptide synthesis, high-throughput screening methods, the use of transgenic animals for modeling of human infections. An important role for the development and selection of efficient adjuvants for peptide immunogens is noted. The review contains examples of the developments of synthetic peptide vaccines against three infectious diseases (malaria, hepatitis C, and foot-and-mouth disease).     

Different applications of virus‐like particles in biology and medicine: Vaccination and delivery systems

Virus‐like particles (VLPs) mimic the whole construct of virus particles devoid of viral genome as used in subunit vaccine design. VLPs can elicit efficient protective immunity as direct immunogens compared to soluble antigens co‐administered with adjuvants in several booster injections. Up to now, several prokaryotic and eukaryotic systems such as insect, yeast, plant, and E. coli were used to express recombinant proteins, especially for VLP production. Recent studies are also generating VLPs in plants using different transient expression vectors for edible vaccines. VLPs and viral particles have been applied for different functions such as gene therapy, vaccination, nanotechnology, and diagnostics. Herein, we describe VLP production in different systems as well as its applications in biology and medicine. © 2015 Wiley Periodicals, Inc. Biopolymers 105: 113–132, 2016.     

DNA-vaccination via tattooing induces stronger humoral and cellular immune responses than intramuscular delivery supported by molecular adjuvants

Tattooing is one of a number of DNA delivery methods which results in an efficient expression of an introduced gene in the epidermal and dermal layers of the skin. The tattoo procedure causes many minor mechanical injuries followed by hemorrhage, necrosis, inflammation and regeneration of the skin and thus non-specifically stimulates the immune system. DNA vaccines delivered by tattooing have been shown to induce higher specific humoral and cellular immune responses than intramuscularly injected DNA. In this study, we focused on the comparison of DNA immunization protocols using different routes of administrations of DNA (intradermal tattoo versus intramuscular injection) and molecular adjuvants (cardiotoxin pre-treatment or GM-CSF DNA co-delivery). For this comparison we used the major capsid protein L1 of human papillomavirus type 16 as a model antigen. L1-specific immune responses were detected after three and four immunizations with 50 μg plasmid DNA. Cardiotoxin pretreatment or GM-CSF DNA co-delivery substantially enhanced the efficacy of DNA vaccine delivered intramuscularly by needle injection but had virtually no effect on the intradermal tattoo vaccination. The promoting effect of both adjuvants was more pronounced after three rather than four immunizations. However, three DNA tattoo immunizations without any adjuvant induced significantly higher L1-specific humoral immune responses than three or even four intramuscular DNA injections supported by molecular adjuvants. Tattooing also elicited significantly higher L1-specific cellular immune responses than intramuscularly delivered DNA in combination with adjuvants. In addition, the lymphocytes of mice treated with the tattoo device proliferated more strongly after mitogen stimulation suggesting the presence of inflammatory responses after tattooing. The tattoo delivery of DNA is a cost-effective method that may be used in laboratory conditions when more rapid and more robust immune responses are required.     

Immunogens Modeling a Fusion-Intermediate Conformation of gp41 Elicit Antibodies to the Membrane Proximal External Region of the HIV Envelope Glycoprotein

The membrane proximal external region (MPER) of the gp41 subunit of the HIV-1 envelope glycoprotein (Env) contains determinants for broadly neutralizing antibodies and has remained an important focus of vaccine design. However, creating an immunogen that elicits broadly neutralizing antibodies to this region has proven difficult in part due to the relative inaccessibility of the MPER in the native conformation of Env. Here, we describe the antigenicity and immunogenicity of a panel of oligomeric gp41 immunogens designed to model a fusion-intermediate conformation of Env in order to enhance MPER exposure in a relevant conformation. The immunogens contain segments of the gp41 N- and C-heptad repeats to mimic a trapped intermediate, followed by the MPER, with variations that include different N-heptad lengths, insertion of extra epitopes, and varying C-termini. These well-characterized immunogens were evaluated in two different immunization protocols involving gp41 and gp140 proteins, gp41 and gp160 DNA primes, and different immunization schedules and adjuvants. We found that the immunogens designed to reduce extension of helical structure into the MPER elicited the highest MPER antibody binding titers, but these antibodies lacked neutralizing activity. The gp41 protein immunogens also elicited higher MPER titers than the gp140 protein immunogen. In prime-boost studies, the best MPER responses were seen in the groups that received DNA priming with gp41 vectors followed by gp41 protein boosts. Finally, although titers to the entire protein immunogen were similar in the two immunization protocols, MPER-specific titers differed, suggesting that the immunization route, schedule, dose, or adjuvant may differentially influence MPER immunogenicity. These findings inform the design of future MPER immunogens and immunization protocols.     

The choice of adjuvants in Mycoplasma vaccines

The use of adjuvants in vaccine production is an important aspect of potent vaccines. This investigation was concerned with finding the most efficient adjuvants for use in Mycoplasma vaccines produced in Nigeria. Four different vaccines were produced from the Gladysdale strain of Mycoplasma mycoides subspecies mycoides. They differed depending on the type of adjuvants used. Each vaccine was used to vaccinate eight cattle using a dose of 1 ml. Two other groups of eight cattle were used as controls. One of the two groups received 1 ml dose of inactivated Gladysdale vaccine without adjuvant while the second group received 1 ml dose of saline. The number of cattle that had the peak complement fixing (CF) antibody titres of 1/80 in each group of cattle was four for vaccine containing aluminium hydroxide gel, eight for vaccine containing liquid paraffin, one for vaccine containing sodium alginate and one for vaccine without adjuvant. Seven cattle from the group vaccinated with vaccine containing Freund's incomplete adjuvant had peak CF antibody titres of 1/80 or higher. The two groups vaccinated with vaccine containing liquid paraffin and Freund's incomplete adjuvant survived challenge at 6 months post vaccination. Freund's incomplete adjuvant and liquid paraffin containing 10% Arlacel A are the most efficient adjuvants.     

Peptides delivered by immunostimulating reconstituted influenza virosomes.

Vaccines have been well accepted and used effectively for more than 100 years. Traditional vaccines are generally composed of whole inactivated or attenuated microorganisms that have lost their disease-causing properties. These classical prophylactic live vaccines evoke protective immune responses, but have often been associated with an unfavorable safety profile, as observed, for example, for smallpox and polio myelitis vaccines [1,2]. First improvements were subunit vaccines that do not focus on attenuation of whole organisms but concentrate on particular proteins. These vaccines are able to generate protective immune responses (e.g. diphtheria, tetanus, pertussis)3. However, next generation vaccines should focus on specific antigens (e.g. proteins, peptides), since the requirements by regulatory authorities to crude biological material are becoming more stringent over time. An increasing number of such antigens capable of inducing protective humoral or cellular immune responses have been identified in the last few years. But most of these are weak immunogens. This reemphasizes the need for adjuvants to promote a potent immune response and also for delivery antigens to the immune system in an appropriate way (carrier capability). Here we review a new approach for prophylactic and therapeutic vaccines, which focuses on the induction of highly specific immune responses directed against antigen-derived peptides using a suitable carrier system.