应用植物表达系统生产疫苗的研究概况

随着重组DNA技术、植物转基因技术和分子植物病毒学的迅猛发展,使得应用植物作为疫苗抗原的表达载体成为可能。本对应用转基因植物和基于植物的病毒载体两种植物表达系统生产疫苗抗原的研究概况进行了评述。     

利用转基因植物生产可食疫苗

主要对转基因植物可食疫苗的转化和表达技术及其优越性和种类进行总结,并对该技术的发展趋势进行了展望。     

利用转基因植物生产口服疫苗的研究进展

人和动物的许多疾病是通过病原微生物与消化道、呼吸道和生殖道的表面粘膜接触而引起的〔1〕,如通过消化道传染的疾病有乙型肝炎、伤寒、霍乱、痢疾和胃肠炎等。在发展中国家 ,因为缺少资金购买或生产昂贵的疫苗和医疗设备 ,患病情况尤为严重。因此 ,急需生产价格低廉且不需要复杂医疗设备的口服疫苗。目前 ,利用转基因植物生产口服疫苗越来越受到重视〔1～ 12〕。这种技术已较成熟 ,主要涉及到口服疫苗抗原的选择、目标植物的选择、表达载体的构建、转化和检测及动物和人体试验。1　口服疫苗抗原的选择利用转基因植物生产的口服疫苗抗原一…     

谈谈植物疫苗问题

本文对植物疫苗的研究现状和问题进行了介绍和讨论。     

转基因植物表达HIV疫苗的研究进展

转基因植物成本低廉,表达的外源蛋白安全性高,易于大规模生产。建立HIV疫苗的植物表达系统,开辟了HIV疫苗研制的新领域。本综述了转基因植物表达HIV疫苗过程中目的基因的选择与修饰、载体的构建、受体植物的选定、目的基因的表达、优势与安全性等研究进展。     

生产重组蛋白的植物表达系统

作为高附加值重组蛋白生产平台,由于在成本和安全性方面的优势,植物已成为继微生物、哺乳动物等表达系统之后,获得广泛认同的极具潜力的蛋白表达系统。植物表达系统主要包括转基因植株,叶绿体转化植物,瞬时表达系统,细胞悬浮培养。综述了这些表达系统生产重组蛋白的产量和质量,尤其是各种表达系统的优缺点。     

蚕杆状病毒载体表达系统及其在动物疫苗生产中的应用

就杆状病毒的分子生物学特性及其作为表达载体以蚕体表达外源基因的特点和优点进行综述,并对该表达系统在动物疫苗生产中的应用作一简要概述,。     

用转基因植物生产基因工程疫苗

用转基因植物生产外源蛋白质产品是一个有吸引力的廉价生产系统,它有可能替代外源蛋白质的发酵生产系统。通过外源基因的瞬时表达或稳定表达方式,多种疫苗已在植物中产生,在植物中表达的抗原保持了它自身的免疫原性,植物在这方面的应用具有独特的优点。     

利用植物系统表达重组蛋白的的研究进展

植物是可以生产不同生物药剂品的成本低廉的生物反应器,综述了稳定转化系统、瞬时表达系统以及叶绿体基因组转化方法,这三种不同的植物表达系统的特点和研究现状,并对其存在的问题及未来的前景进行了分析。     

利用转基因植物生产生物药

传统的生物技术常利用哺乳动物细胞、细菌和真菌培养作为转基因系统 ,来生产生物药。鉴于今后对生物药例如治疗贫血的红细胞生成素和治疗糖尿病的胰岛素 ,以及通过基因组研究发现的新的药用蛋白的需求量均将大量增加 ,故有必要利用另一种转基因系统来生产价格既低廉 ,使用又安全的重组生物药。认为适合于上述目的者为高等植物。利用转基因植物来生产重组药用蛋白和肽有很多优点 ,例如 :(1)植物容易转化 ;(2 )使用安全 ,因植物不是人类病原体的宿主 ,故与动物细胞培养比较 ,利用重组植物生产的生物药不论提纯与否 ,都不大有可能污染人类病原…     

Plants as biofactories

Cell substrates are a key component of successful vaccine development and throughout the last several decades there has been a dramatic increase in the types of cells available for vaccine production. Nevertheless, there is a continued demand for new and innovative approaches for vaccine development and manufacturing. Recent developments involving cells of insect and plant origin are attracting considerable scientific interest. Here we review vaccine antigen production in plant-based systems as was presented by Dr. Vidadi Yusibov of Fraunhofer USA Center for Molecular Biotechnology at the IABS International Scientific Workshop on NEW CELLS FOR NEW VACCINES II that was held in Wilmington, Delaware on September 17–19, 2007.     

昆虫杆状病毒表达载体系统在疫苗研究中的应用进展

昆虫杆状病毒表达载体系统(Baculovirus expression vector system,BEVS)已成功应用于多种蛋白的表达,并为疫苗开发提供了充足的原材料。相比其他表达系统,BEVS具有许多优势:杆状病毒专一寄生于无脊椎动物,安全性高;重组蛋白表达水平高;可对重组蛋白进行正确折叠和翻译后修饰,获得具有生物活性的蛋白;适应于多基因表达如病毒样颗粒(Virus-like particle)的复杂设计;适用于大规模无血清培养等。为了更好地理解BEVS在疫苗研究中的应用前景,文中将从BEVS的发展及其在疫苗研究中的应用等方面进行综述。     

Biodistribution and expression of targeted fusion anti-caries DNA vaccine pGJA-P/VAX in mice

OBJECTIVE: To study the biodistribution and expression of anti-caries DNA vaccine pGJA-P/VAX in mice models following intranasal (i.n.) and intramuscular (i.m.) immunization. METHODS: pGJA-P/VAX and pVAX1 were administrated i.n. and i.m. to mice, respectively. Blood and organs were harvested and then genomic DNA was extracted. mRNA in the original tissues and draining lymph nodes was isolated. The in vivo fate of plasmid DNA in these samples was evaluated with real-time quantitative polymerase chain reaction (PCR). RESULTS: The ratio of the area under the curve (AUC) and maximum concentration (Cmax) after i.n. immunization were higher than that after i.m. injection. At 1 day post i.n. immunization, the localization of plasmid to the heart, spleen and lung was notable compared with that to the nasal mucosa. At 2 days, for i.n. immunization, the concentration value of plasmid in the mucosa-associated lymphoid tissues such as lung, kidney and gastrointestine had increased and was greater than that via i.m. injection. At later time points, the plasmid was primarily in the original tissues and draining lymph nodes. mRNA expression was observed in the topical tissues up to 7 days and in the draining lymph nodes for at least 3 days. CONCLUSIONS: These results indicate that the distribution of pGJA-P/VAX was widespread in vivo, and the expression was significant in the inoculation tissue and draining lymph nodes after i.n. immunization and i.m. injection. This data provides evidence which supports the long-term immune responses and the basis of further research on safety issues.     

Improved understanding of gene expression regulation using systems biology

This article reviews the current state of systems biology approaches, including the experimental tools used to generate ‘omic’ data and computational frameworks to interpret this data. Through illustrative examples, systems biology approaches to understand gene expression and gene expression regulation are discussed. Some of the challenges facing this field and the future opportunities in the systems biology era are highlighted.     

Detailed functional characterization of glycosylated and nonglycosylated variants of malaria vaccine candidate PfAMA1 produced in Nicotiana benthamiana and analysis of growth inhibitory responses in rabbits

One of the most promising malaria vaccine candidate antigens is the Plasmodium falciparum apical membrane antigen 1 (PfAMA1). Several studies have shown that this blood‐stage antigen can induce strong parasite growth inhibitory antibody responses. PfAMA1 contains up to six recognition sites for N‐linked glycosylation, a post‐translational modification that is absent in P. falciparum. To prevent any potential negative impact of N‐glycosylation, the recognition sites have been knocked out in most PfAMA1 variants expressed in eukaryotic hosts. However, N‐linked glycosylation may increase efficacy by improving immunogenicity and/or focusing the response towards relevant epitopes by glycan masking. We describe the production of glycosylated and nonglycosylated PfAMA1 in Nicotiana benthamiana and its detailed characterization in terms of yield, integrity and protective efficacy. Both PfAMA1 variants accumulated to high levels (>510 μg/g fresh leaf weight) after transient expression, and high‐mannose‐type N‐glycans were confirmed for the glycosylated variant. No significant differences between the N. benthamiana and Pichia pastoris PfAMA1 variants were detected in conformation‐sensitive ligand‐binding studies. Specific titres of >2 × 10⁶ were induced in rabbits, and strong reactivity with P. falciparum schizonts was observed in immunofluorescence assays, as well as up to 100% parasite growth inhibition for both variants, with IC₅₀ values of ~35 μg/mL. Competition assays indicated that a number of epitopes were shielded from immune recognition by N‐glycans, warranting further studies to determine how glycosylation can be used for the directed targeting of immune responses. These results highlight the potential of plant transient expression systems as a production platform for vaccine candidates.     

Benefits and risks of antibody and vaccine production in transgenic plants

Phytopharming, the production of protein biologicals in recombinant plant systems, has shown great promise in studies performed over the past 13 years. A secretory antibody purified from transgenic tobacco was tested successfully in humans, and prevented bacterial re-colonization after topical application in the mouth. Rapid production of patient-tailored anti-lymphoma antibodies in recombinant Tobamovirus-infected tobacco may provide effective cancer therapy. Many different candidate vaccines from bacterial and viral sources have been expressed in transgenic plants, and three human clinical trials with oral delivery of transgenic plant tissues have shown exciting results. The use of crop plants with agricultural practice could allow cheap production of valuable proteins, while providing enhanced safety by avoidance of animal viruses or other contaminants. However development of this technology must carefully consider the means to ensure the separation of food and medicinal products when crop plants are used for phytopharming.     

植物中的P-ATP酶

P-ATP酶是指位于质膜上、由ATP驱动的一类经历磷酸化的阳离子泵,它普遍存在于各种生物中,广泛参与植物的离子运输、细胞信号转导、细胞膜形态建成等,与植物的离子养分吸收运输、逆境抗性等密切相关。     

转基因植物表达重组蛋白的研究进展

植物表达系统的一些潜在优点 ,如重组蛋白的高积累水平 ,糖基化 ,细胞内的定位和自然储藏的稳定性是目前植物生产重组蛋白系统研究成为热点的主要原因 .在研究和选择转基因植物表达系统的过程中 ,转化 ,转化后 ,翻译 ,翻译后等环节都会影响到最终产物的数量和质量 ,因此应该了解基因表达的规律 ,以制定植物生产重组蛋白合适的策略 ,重组蛋白积累水平是关键 ,但其它因素如植物的选择 ,转基因植物的处理 ,下游加工等同样重要 .某些情形下 ,仅下游加工的成本一项就影响到特定植物表达系统的实际应用价值 .     

Plants as bioreactors for the production of vaccine antigens

Plants have been identified as promising expression systems for commercial production of vaccine antigens. In phase I clinical trials several plant-derived vaccine antigens have been found to be safe and induce sufficiently high immune response. Thus, transgenic plants, including edible plant parts are suggested as excellent alternatives for the production of vaccines and economic scale-up through cultivation. Improved understanding of plant molecular biology and consequent refinement in the genetic engineering techniques have led to designing approaches for high level expression of vaccine antigens in plants. During the last decade, several efficient plant-based expression systems have been examined and more than 100 recombinant proteins including plant-derived vaccine antigens have been expressed in different plant tissues. Estimates suggest that it may become possible to obtain antigen sufficient for vaccinating millions of individuals from one acre crop by expressing the antigen in seeds of an edible legume, like peanut or soybean. In the near future, a plethora of protein products, developed through ‘naturalized bioreactors’ may reach market. Efforts for further improvements in these technologies need to be directed mainly towards validation and applicability of plant-based standardized mucosal and edible vaccines, regulatory pharmacology, formulations and the development of commercially viable GLP protocols. This article reviews the current status of developments in the area of use of plants for the development of vaccine antigens.     

A vaccine strategy utilizing a combination of three different chimeric vectors which share specific vaccine antigens

A large number of recombinant of viral and bacterial systems have been engineered as vectors to express foreign genes for vaccination and/or gene therapy. A common problem is the immune response to the vector itself. The presence of anti-vector immune responses may preclude sufficient 'priming' or immunogenicity if pre-existing immune responses are present, or they may impair optimal 'boosting' upon repeated immunization or delivery with the same vector. To circumvent this problem we developed a strategy using different chimeric vectors which share only the expression of common specific antigens desired for immunization. This approach not only has the advantage of avoiding increased anti-vector responses, but allows the use of combinations of vectors which could subsequently present the same or related antigen differently to the immune system as well as at alternative sites to induce the optimal type of immunity against the pathogen of interest.