WO2017113786A1 - 突变的病毒、其制备方法和应用 - Google Patents
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- the plasmid expressing a tRNA and a tRNA synthetase that specifically recognizes an unnatural amino acid such as Lys-diazirine and Lys-azido is a pACYC-tRNA/PylRS plasmid which has been deposited in a general microbial species collection management in China. Center, the deposit date is June 14, 2011. It has been disclosed in the patent of Chinese Patent No. 201210214451.6.
- a person skilled in the art can determine the site suitable for introducing the TAG mutation in the coding gene of each protein of the influenza virus by various methods, for example, analyzing the amino acid conservation in the corresponding protein of the influenza virus by bioinformatics means, and then determining the introduction of the TAG. The location.
- the inventors in order to improve the efficiency of influenza virus production, and in future industrial production, the inventors established a stable expression of tRNA (tRNA Pyl ) and pyrrolysyl-tRNA synthetase (tRNA Pyl ).
- Mammalian stable cell line HEK293-PYL Mammalian stable cell line.
- the mammalian stable cell line also solves the shortcomings of the conventional use of chicken embryo propagation virus to cause adverse reactions such as human allergy.
- the present invention provides a mutant influenza virus which can be prepared in mammalian cells, introducing an amber stop codon UAG and an unnatural amino acid, respectively, in the genome and corresponding proteins.
- the mutated virus according to item 1 which is an influenza virus, and the nucleic acid encoding the at least one protein selected from the group consisting of PA, PB1, PB2, NP, NA, HA, NS or M protein in the influenza virus at the termination password
- One or more codons upstream of the child contain one or more UAG codons.
- the mutated influenza virus of item 3 wherein the nucleic acid encoding at least two, three or four proteins selected from the group consisting of PA, PB1, PB2, NP, NA, HA, NS or M proteins is upstream of the stop codon
- One or more codons contain one or more UAG codons.
- influenza virus according to item 3, wherein the influenza virus is selected from the following Tables 2a), 2b), 2c), 2d), 2e), 2f), 2g), 2h) and 2i) UAG codons are included at one or more of the listed gene loci.
- n is any position within the qth position upstream of the start codon of the protein to the qth position upstream of the natural stop codon, wherein p and q are independently selected from 1, 2, 3, 4, 5, 6 , 7, 8, 9, 10, 15, 20, 30, 40, 50, 60, 70, 80, 90, 100, 125, 150, 200, 250, 300, 350, 400, 450, 500, 600, 700 Or until the length of its full length amino acid sequence is minus one.
- the mutated virus of item 11 which is an influenza virus, and comprises an unnatural amino acid at R266 on PA, R52 on PB1, K33 on PB2, and D101 on NP.
- the amino acid sequence of the unmutated PB1 is identical to the amino acid sequence encoded by the nucleic acid shown in SEQ ID NO:
- the amino acid sequence of the unmutated NA is the same as the amino acid sequence encoded by the nucleic acid shown in SEQ ID NO: 7,
- the mutated virus according to item 14 which is a WSN-RNP-TAG comprising a UAG codon at the coding codon of R266 of the PA protein, R52 of the PB1 protein, K33 of the PB2 protein and D101 of the NP protein.
- the animal cell line is selected from the group consisting of a mammalian cell line, an avian animal cell line, a hamster cell line, and the like, preferably selected from the group consisting of 293 cells, 293T cells, Vero cells, A549 cells, Hela cells, CHO cells, MDCK cells, sf9 cells, and the like.
- step (3) is a plasmid in Escherichia coli pACYC-tRNA/PylRS deposited on June 14, 2011 and deposited under CGMCC No:4951. pACYC-tRNA/PylRS.
- Figure 2 Effect of rescue of influenza virus after point mutations on individual proteins of influenza virus. Sites are only representative and are not limited to the sites shown.
- Figure 2G Effect of rescue of influenza virus after point mutation of influenza virus NS protein.
- Figure 5 The yield of influenza virus prepared using the method of our invention is high by qRT-PCR, close to the production of wild-type influenza virus.
- FIG. 9B Detection of anti-HA antibody levels in mice of each group before and after immunization by Elisa assay. The results showed that the antibody content in the mice increased both 14 days after immunization and 21 days after immunization, and the antibody content at 21 days was higher than that at 14 days.
- FIG. 10 Comparison of the efficiency of rescue of mutant influenza virus using the mammalian stable cell line HEK293-PYL, which is a normal mammalian cell 293T and stably expressing tRNA (tRNA Pyl ) and pyrrolysyl-tRNA synthetase (tRNA Pyl ). The results indicate that the use of the HEK293-PYL cell line to rescue mutant influenza viruses is much more efficient than the use of common 293T cells.
- tRNA Pyl a normal mammalian cell 293T and stably expressing tRNA
- tRNA Pyl pyrrolysyl-tRNA synthetase
- tRNA Pyl HEK293-PYL stably expressing tRNA
- tRNA Pyl pyrrolysyl-tRNA synthetase
- Two lentiviral overexpression vectors with puromycin and hygromycin resistance were constructed, respectively carrying an aminoacyl tRNA synthetase and a reporter gene GFP carrying the TAG mutation at the codon 39 coding codon, which was transferred by two rounds of virus.
- HEK-293T cells and puromycin/hygromycin were screened to obtain a stable cell line pylRS/GFP 39TAG .
- the bjmu-zeo-12tRNA vector carrying 12 copy number tRNA and zeomycin resistance was constructed, and the plasmid was linearized and transfected into pylRS/GFP 39TAG , then screened by zeomycin in the presence of unnatural amino acid (UAA), and finally separated.
- UAA unnatural amino acid
- the inventors also examined the yield of the prepared mutant influenza virus WSN-RNP-tag.
- the yield of the prepared mutant influenza virus was substantially identical to that of the wild type (Fig. 5).
- the antigen red blood cell agglutination titer of wild-type and mutant WSN-RNP-TAG viruses it is indicated that the antigenic red blood cell agglutination titers of the mutant influenza virus and the wild-type virus are substantially identical (Fig. 6), further illustrating the preparation.
- the yield of the mutant influenza virus is high.
- Specific experimental procedures can refer to the experimental procedure of qRT-PCR and the antigen red blood cell agglutination titer assay method of virus.
- mice were weighed, and 5 of each group of mice were selected, and their serum was taken (the blood collection volume was 20-40 ul, which prevented the mice from losing blood.
- the collected serum was at -80 °C frozen storage).
- the first group was inoculated with the virus solution of No. 1 tube (ingredient: PBS);
- the third group was inoculated with the virus solution of the No. 3 tube (ingredient: 1 ⁇ lethal dose of WSN-RNP-tag);
- the fourth group was inoculated with the virus solution of the No. 4 tube (ingredient: 5 times lethal dose of WSN-RNP-tag);
- the sixth group was inoculated with the virus solution of the No. 6 tube (ingredient: 1-fold lethal dose of inactivated WSN);
- the seventh group was inoculated with the virus solution of the No. 7 tube (ingredient: 5 times lethal dose of inactivated WSN);
- the eighth group was inoculated with the virus solution of tube No. 8 (ingredient: 10 times lethal dose of inactivated WSN);
- mice were taken from each group of mice, their lung tissues were taken, and after homogenization, the supernatant was collected (can be stored at -80 ° C for cryopreservation). The remaining mice continue to be observed.
- the antibody content of anti-NA and NP proteins in mice was measured using the Elisa method. Specifically, the protein NA or NP was diluted with a coating solution, 30 ng/100 ul. A 96-well plate for Elisa was coated with the above dilution at 4 ° C overnight. After washing with Elisa washing solution, the mixture was blocked with a washing solution containing 3% BSA for 1 h at 37 ° C, and then the mouse serum was diluted with a washing solution of 0.5% BSA, added to the corresponding air, incubated at 37 ° C for 1 h, and the supernatant was discarded. Wash 5 times with washing solution.
- Method of inoculation of the virus anesthesia, nasal inoculation of 50 ul of virus solution into the nasal cavity of the mouse.
- the median lethal dose LD50 was 10,000 viral particles per 50 ul.
- the doubling dose is 10*LD50 and the 10-fold lethal dose is 100*LD50.
- the third group was inoculated with the virus solution of the No. 3 tube (ingredient: 10-fold lethal dose of inactivated WSN);
- Elisa was used to detect the production of IgA. Specifically, the purified WSN virus was diluted with a coating solution, 0.5 ug/100 ul. A 96-well plate for Elisa was coated with the above dilution at 4 ° C overnight. After washing with Elisa washing solution, the mixture was blocked with a washing solution containing 3% BSA for 1 h at 37 ° C, and then the mouse serum was diluted with a washing solution of 0.5% BSA, added to the corresponding air, incubated at 37 ° C for 1 h, and the supernatant was discarded. Wash 5 times with washing solution.
- Test method After three weeks of immunization of the mice, the lung tissues of the mice were taken and the T lymphocytes were isolated. T lymphocytes were stained with anti-mouse CD8a-APC antibody and influenza NP366-374-tetramer-PE. The number (in proportion) of influenza-specific CD8+ T cells was measured by flow cytometry.
- Method please refer to the literature Budimir, N. et al. Heterosubtypic cross-protection induced by whole inactivated influenza virus vaccine in mice: influence of the route of Vaccine administration.Influenza Other Respir Viruses 7,1202-1209 (2013).
- the mutant virus reduces the titer of the progeny virus of the wild type virus, and increasing the titer of the mutant virus or increasing the number of UAG codons in the mutant virus genome enhances the mutant type. Inhibition of viral replication by wild-type virus.
- mice were randomly removed from each group, and lung tissues were taken out to measure the titer of wild-type virus in lung tissues.
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Abstract
Description
Claims (39)
- 一种突变的病毒,其特征在于所述病毒的至少一种蛋白的编码核酸在终止密码子上游的一个或多个密码子处包含一个或多个UAG密码子。
- 权利要求1所述的突变的病毒,其选自下述组中:手足口病毒、柯萨奇病毒、丙肝病毒HCV、乙肝病毒HBV、甲肝病毒、丁型肝炎病毒、戊型肝炎病毒、EB病毒、人乳头瘤病毒HPV、单纯疱疹病毒HSV、巨细胞病毒、水痘-带状疱疹病毒、水泡性口炎病毒、呼吸道合胞病毒RSV、登革病毒、埃博拉病毒、寨卡病毒Zika、SARS、中东呼吸综合征病毒、轮状病毒、狂犬病毒、麻疹病毒、腺病毒、艾滋病毒、脊髓灰质炎病毒、埃可病毒、乙型脑炎病毒、森林脑炎病毒、汉坦病毒、新型肠道病毒、风疹病毒、腮腺炎病毒、副流感病毒、蓝耳病毒、猪瘟病毒、口蹄疫病毒、细小病毒。
- 权利要求1所述的突变的病毒,其是流感病毒,并且述流感病毒中选自PA、PB1、PB2、NP、NA、HA、NS或M蛋白的至少一种蛋白的编码核酸在终止密码子上游的一个或多个密码子处包含一个或多个UAG密码子。
- 权利要求3所述的突变的流感病毒,其中选自PA、PB1、PB2、NP、NA、HA、NS或M蛋白的至少两种、三种或四种蛋白的编码核酸在终止密码子上游的一个或多个密码子处包含一个或多个UAG密码子。
- 权利要求3所述的突变的流感病毒,其中所述PA、PB1、PB2和NP蛋白中两种、三种或四种蛋白的编码核酸在终止密码子上游的一个或多个密码子处包含一个或多个UAG密码子。
- 权利要求3所述的突变的流感病毒,其中所述流感病毒在选自说明书表2a)、2b)、2c)、2d)、2e)、2f)、2g)、2h)和2i)中所列举的一个或多个基因位点处包含UAG密码子。
- 权利要求3所述的突变的流感病毒,其中所述流感病毒在PA蛋白的R266、PB1蛋白的R52、PB2蛋白的K33和/或NP蛋白的D101的编码核酸密码子处含有UAG密码子。
- 如权利要求8所述的突变的病毒,其中所述的非天然氨基酸是位于第n位的Lys-diazirine,其在病毒蛋白中的连接方式如下式所示:其中,由R1到R2的方向为氨基酸序列的N末端到C末端方向,第n位可以是流感病毒的PA、PB1、PB2、NP、NA、HA、NS或M蛋白中任何一种蛋白质氨基酸序列内的任意位置,相应地,R1为第1至第n-1位氨基酸残基,R2为第n+1位至C末端的氨基酸残基,优选地,n是该蛋白起始密码子下游第p-1位到天然终止密码子上游第q位内的任意位置,其中p和q独立地选自1、2、3、4、5、6、7、8、9、10、15、20、30、40、50、60、70、80、90、100、125、150、200、250、300、350、400、450、500、600、700或者直到其全长氨基酸序列长度减去1。
- 如权利要求8所述的突变的病毒,其中所述的非天然氨基酸是位于第n位的Lys-azido,其在病毒蛋白中的连接方式如下式所示:其中,由R1到R2的方向为氨基酸序列的N末端到C末端方向,第n位可以是病毒的PA、PB1、PB2、NP、NA、HA、NS或M蛋白中任何一种蛋白质的氨基酸序列内的任意位置,相应地,R1为第1至第n-1位氨基酸残基,R2为第n+1位至C末端的氨基酸残基,优选地,n是该蛋白起始密码子下游第p-1位到天然终止密码子上游第q位内的任意位置,其中p和q独立地选自1、2、3、4、5、6、7、8、9、10、15、20、30、40、50、60、70、80、90、100、125、150、200、250、300、350、400、450、500、600、700或者直到其全长氨基酸序列长度减去1。
- 权利要求8-10中任一项的突变的病毒,其在所述蛋白质的多个位点处包含非天然氨基酸。
- 如权利要求11所述的突变的病毒,其是流感病毒,并且在其中PA上的R266、PB1上的R52、PB2上的K33、NP上的D101处包含非天然氨基酸。
- 如权利要求1-12中任一项所述的病毒,其中所述病毒的蛋白质编码序列中的正常终止密码子如果是UAG的话,已被突变成UAA。
- 如权利要求3-13中任一项所述的突变的病毒,其是流感病毒,并且,未经突变的PB2的氨基酸序列与SEQ ID NO:2所示的核酸编码的氨基酸序列相同,未经突变的PB1的氨基酸序列与SEQ ID NO:3所示的核酸编码的氨基酸序列相同,未经突变的PA的氨基酸序列与SEQ ID NO:4所示的核酸编码的氨基酸序列相同,未经突变的HA的氨基酸序列与SEQ ID NO:5所示的核酸编码的氨基酸序列相同,未经突变的NP的氨基酸序列与SEQ ID NO:6所示的核酸编码的氨基酸序列相同,未经突变的NA的氨基酸序列与SEQ ID NO:7所示的核酸编码的氨基酸序列相同,未经突变的M的氨基酸序列与SEQ ID NO:8所示的核酸编码的氨基酸序列相同,或未经突变的NS的氨基酸序列与SEQ ID NO:9所示的核酸编码的氨基酸序列相同。
- 根据权利要求14所述的突变的病毒,其是WSN-RNP-TAG,在PA蛋白的R266、PB1蛋白的R52、PB2蛋白的K33和NP蛋白的D101的编码密码子处含有UAG密码子。
- 权利要求3所述的突变的病毒,其是人源的或其它动物源的流感病毒,优选为A、B或C型流感病毒。
- 权利要求3所述的突变的病毒,其中包含有说明书表2a-表2i和表4中所列举的一个或多个突变。
- 一种编码如权利要求1-17中任一项所述的突变的病毒或其经突变的蛋白的核酸分子,其特征在于在天然终止密码子之外,还包含人为引入的密码子UAG。
- 制备权利要求1-17中任一项所述突变的病毒的方法,包括如下步骤:(1)将包含所述一个或多个被突变基因与合适的载体可操作地连接的突变核酸表达载体和包含其它未被突变基因的一个或多个核酸表达载体转染稳定表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA合成酶(tRNAPyl)的动物细胞系,或者与表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA合成酶(tRNAPyl)的质粒共同转染动物细胞系其它未被突变基因的一个或多个核酸表达载体;(2)将被转染的细胞在含有Lys-diazirine或者Lys-azido的培养基中培养;和(3)回收所述突变病毒。
- 权利要求19所述的方法,其中步骤(1)中所述质粒是保藏日为2011年6月14日、保藏号为CGMCC No:4951的大肠埃希氏菌pACYC-tRNA/PylRS中的质粒pACYC-tRNA/PylRS。
- 权利要求19所述的方法,其中所述动物细胞系选自哺乳动物细胞系、禽类动物细胞系、仓鼠细胞系等,优选选自293细胞、293T细胞、Vero细胞、A549细胞、Hela细胞、CHO细胞、MDCK细胞、sf9细胞。
- 权利要求19所述的方法,其中所述稳定表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA 合成酶(tRNAPyl)的动物细胞系是以保藏号CGMCC No:11592于2015年11月17日被保藏在CGMCC的HEK293-PYL。
- 筛选减毒病毒的方法,包括步骤:(1)基因突变:通过基因工程方法将病毒基因组中一个或多个基因的一个或多个选定的密码子突变为TAG密码子,得到一个或多个被突变的基因;(2)表达载体构建:将(1)得到的一个或多个被突变的基因与合适的载体可操作地连接,得到突变基因表达载体;(3)将步骤(2)中得到的突变基因表达载体和包含其它未被突变基因的一个或多个基因表达载体转染稳定表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA合成酶(tRNAPyl)的动物细胞系,或者与表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA合成酶(tRNAPyl)的质粒共同转染动物细胞系;(4)将被转染的细胞在含有Lys-diazirine或者Lys-azido的培养基中培养,收集培养基上清液,采用包含Lys-diazirine或者Lys-azido的培养板对上清液进行病毒克隆的非天然氨基酸依赖性的检测;和(5)将维持了对Lys-diazirine或者Lys-azido非天然氨基酸依赖性的突变体鉴定为减毒的病毒。
- 权利要求23的方法,其在步骤(5)之后还包括:(6)将(5)中定点突变成功的候选物的突变位点进行组合,再次包装病毒,使得包装出的病毒在多个基因片段上均引入了UAG,在对应的多个蛋白上均引入了非天然氨基酸,然后再次考察包装产物对非天然氨基酸的依赖性,保留经过长期传代仍旧维持着对非天然氨基酸依赖性的组合突变体设定为最优的定点突变成功候选物。
- 权利要求24的方法,其中在步骤(6)之后还进一步包括:(7)对(6)中获得的突变型病毒进行安全性,鉴定出安全的病毒。
- 权利要求23所述的方法,其中所述病毒选自下述组中:手足口病毒、柯萨奇病毒、丙肝病毒HCV、乙肝病毒HBV、甲肝病毒、丁型肝炎病毒、戊型肝炎病毒、EB病毒、人乳头瘤病毒HPV、单纯疱疹病毒HSV、巨细胞病毒、水痘-带状疱疹病毒、水泡性口炎病毒、呼吸道合胞病毒RSV、登革病毒、埃博拉病毒、寨卡病毒Zika、SARS、中东呼吸综合征病毒、轮状病毒、狂犬病毒、麻疹病毒、腺病毒、艾滋病毒、脊髓灰质炎病毒、埃可病毒、乙型脑炎病毒、森林脑炎病毒、汉坦病毒、新型肠道病毒、风疹病毒、腮腺炎病毒、副流感病毒、蓝耳病毒、猪瘟病毒、口蹄疫病毒、细小病毒。
- 权利要求23所述的方法,其中所述一个或多个选定的密码子独立地位于编码序列第n位,n是该蛋白起始密码子下游第p-1位到天然终止密码子上游第q位内的任意位置,其中p和q独立地选自1、2、3、4、5、6、7、8、9、10、15、20、30、40、50、60、70、80、90、100、125、150、200、250、300、350、400、450、500、600、700或者直到其全长氨基酸序列长度减去1。
- 权利要求23所述的方法,其是流感病毒。
- 权利要求23所述的方法,其中步骤(3)中所述质粒是保藏日为2011年6月14日、保藏号为CGMCC No:4951的大肠埃希氏菌pACYC-tRNA/PylRS中的质粒pACYC-tRNA/PylRS。
- 权利要求23所述的方法,其中所述动物细胞系选自细胞系哺乳动物、禽类动物细胞系、仓鼠细胞系等,优选选自293细胞、293T细胞、Vero细胞、A549细胞、Hela细胞、CHO 细胞、MDCK细胞、sf9细胞。。
- 权利要求23所述的方法,其中所述稳定表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA合成酶(tRNAPyl)的动物细胞系是以保藏号CGMCC No:11592于2015年11月17日被保藏在CGMCC的HEK293-PYL。
- 权利要求23所述的方法,其特征在于重复突变、包装和筛选步骤。
- 权利要求23-32中任一项所述的方法,其中所述流感病毒是A型、B型或C型流感病毒。
- 含有有效量的权利要求1-17中任一项所述的突变的流感病毒的组合物。
- 含有有效量的权利要求1-17中任一项所述的突变的流感病毒的疫苗。
- 药物组合物,其中含有有效量的权利要求1-17中任一项所述的突变的流感病毒,以及药学上可以接受的赋形剂。
- 权利要求1-17中任一项所述的突变的流感病毒在制备减毒活疫苗或者制备预防或治疗流感病毒感染相关药物中的用途。
- 权利要求1-17中任一项所述的突变的流感病毒在预防和治疗流感病毒感染中的用途。
- 可稳定表达tRNA(tRNAPyl)和吡咯赖氨酰-tRNA合成酶(tRNAPyl)的哺乳动物稳定细胞系,其为HEK293-PYL,保藏日为2015年11月17日,其保藏号为CGMCC No:11592。
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Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN110743007A (zh) * | 2019-09-10 | 2020-02-04 | 安徽智飞龙科马生物制药有限公司 | 一种联合疫苗及其制备方法、应用 |
CN110846311A (zh) * | 2018-08-20 | 2020-02-28 | 北京大学 | 利用抑制性tRNA系统制备PTC稳定细胞系及应用 |
WO2020181837A1 (en) * | 2019-03-13 | 2020-09-17 | Zhejiang Senwei Biopharmaceutical Development Co., Ltd. | Method for rescuing influenza virus and composition therefor |
CN116904403A (zh) * | 2023-09-13 | 2023-10-20 | 北京民海生物科技有限公司 | 一种腮腺炎病毒突变体及其应用 |
Families Citing this family (16)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN107012121B (zh) * | 2016-01-27 | 2021-01-26 | 北京大学 | 携带正交tRNA/氨酰tRNA合成酶的稳定细胞系的构建 |
CN109295100A (zh) * | 2017-07-25 | 2019-02-01 | 北京大学 | 携带正交tRNA/氨酰tRNA合成酶的稳定细胞系的构建 |
CN107964035B (zh) * | 2017-12-06 | 2020-07-24 | 中国科学院微生物研究所 | 一种用于流感病毒的有限复制型黏膜免疫疫苗 |
CN109180680B (zh) * | 2018-08-01 | 2020-05-08 | 苏州大学 | 一种紫外光触发交联型近红外分子探针及其制备方法与应用 |
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WO2021004389A1 (zh) * | 2019-07-05 | 2021-01-14 | 司龙龙 | 蛋白水解靶向病毒、其活疫苗及其制备方法和用途 |
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CN113502274A (zh) * | 2021-06-21 | 2021-10-15 | 武汉大学 | 一种表达人乳头瘤病毒衣壳蛋白l2的重组流感病毒株及其制备方法与应用 |
CN115074330B (zh) * | 2022-04-19 | 2023-12-29 | 中国医学科学院医学生物学研究所 | 一种基于遗传密码子扩展技术的改造Vero细胞系 |
WO2024040453A1 (zh) * | 2022-08-24 | 2024-02-29 | 北京大学宁波海洋药物研究院 | 突变的流感病毒、药物组合物及用途 |
Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN104592364A (zh) * | 2013-10-30 | 2015-05-06 | 北京大学 | 定点突变和定点修饰的腺相关病毒、其制备方法及应用 |
Family Cites Families (11)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DK1456360T3 (en) * | 2001-04-19 | 2015-08-31 | Scripps Research Inst | Methods and Composition for Preparation of Orthogonal TRNA-Aminoacyl-TRNA Synthetase Pairs |
CA2454397A1 (en) * | 2001-07-20 | 2003-06-05 | The University Of Georgia Research Foundation, Inc. | Attenuated rabies virus with nucleoprotein mutation at the phosphorylation site for vaccination against rabies and gene therapy in the cns |
BRPI0906388A2 (pt) * | 2008-02-08 | 2015-07-07 | Scripps Research Inst | Quebra de tolerância imunológica com um aminoácido não natural geneticamente codificado |
MX348657B (es) * | 2008-09-26 | 2017-06-21 | Ambrx Inc | Microorganismos y vacunas dependientes de replicacion de aminoacidos no naturales. |
WO2011044561A1 (en) * | 2009-10-09 | 2011-04-14 | The Research Foundation Of State University Of New York | Attenuated influenza viruses and vaccines |
CN102838663B (zh) * | 2011-06-23 | 2014-06-11 | 北京大学 | 定点突变和定点修饰的病毒膜蛋白、其制备方法及其应用 |
US9732367B2 (en) * | 2012-09-24 | 2017-08-15 | Medimmune Limited | Cell lines |
CN102964432B (zh) * | 2012-11-22 | 2014-02-05 | 中国科学院微生物研究所 | 抑制a型流感病毒m1蛋白发生磷酸化的方法及其应用 |
EP3087088B1 (en) * | 2013-12-23 | 2022-05-25 | University Of Rochester | Methods and compositions for ribosomal synthesis of macrocyclic peptides |
WO2015120459A2 (en) * | 2014-02-10 | 2015-08-13 | Nutech Ventures | Live, attenuated vaccines and methods of making and using |
MA40773A (fr) * | 2014-10-02 | 2017-08-08 | Vertex Pharma | Variants du virus influenza a |
-
2015
- 2015-12-31 CN CN201511029463.1A patent/CN106929482A/zh active Pending
-
2016
- 2016-08-02 CA CA3010015A patent/CA3010015A1/en not_active Abandoned
- 2016-08-02 EP EP16880581.0A patent/EP3399029A4/en active Pending
- 2016-08-02 AU AU2016383201A patent/AU2016383201A1/en not_active Abandoned
- 2016-08-02 BR BR112018013168-7A patent/BR112018013168A2/zh not_active IP Right Cessation
- 2016-08-02 JP JP2018534937A patent/JP2019500051A/ja active Pending
- 2016-08-02 WO PCT/CN2016/092778 patent/WO2017113786A1/zh active Application Filing
- 2016-08-02 CN CN201680076047.3A patent/CN109563492B/zh active Active
- 2016-08-02 US US16/066,283 patent/US11510974B2/en active Active
-
2018
- 2018-06-25 IL IL260258A patent/IL260258A/en unknown
- 2018-06-27 CO CONC2018/0006706A patent/CO2018006706A2/es unknown
- 2018-06-28 ZA ZA2018/04362A patent/ZA201804362B/en unknown
Patent Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN104592364A (zh) * | 2013-10-30 | 2015-05-06 | 北京大学 | 定点突变和定点修饰的腺相关病毒、其制备方法及应用 |
Non-Patent Citations (8)
Title |
---|
BUDIMIR, N. ET AL.: "Heterosubtypic cross-protection induced by whole inactivated influenza virus vaccine in mice: influence of the route of vaccine administration", INFLUENZA OTHER RESPIR VIRUSES, vol. 7, 2013, pages 1202 - 1209 |
J. W. CHIN; P. G. SCHULTZ, CHEMBIOCHEM, vol. 11, 2002, pages 1135 - 1137 |
J.W. CHIN ET AL., JOURNAL OF THE AMERICAN CHEMICAL SOCIETY, vol. 124, 2002, pages 9026 - 9027 |
L. WANG ET AL., SCIENCE, vol. 292, 2001, pages 498 - 500 |
NEUMANN, G. ET AL., PROC. NATL. ACAD. SCI. USA, vol. 96, 1999, pages 9345 - 50 |
See also references of EP3399029A4 |
TAKAHIRO, H. ET AL.: "Incorporation of Non- Natural Amino Acids into Proteins", CURR OPIN CHEM BIOL., vol. 6, no. 6, 1 December 2002 (2002-12-01), XP008099492, ISSN: 1367-5931 * |
YANG XIA ET AL.: "Genetic Code Expanding Technology", CHEMISTRY OF LIFE, vol. 24, no. 6, 15 December 2004 (2004-12-15), pages 512 - 513, XP009511786, ISSN: 1000-1336 * |
Cited By (6)
Publication number | Priority date | Publication date | Assignee | Title |
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CN110846311A (zh) * | 2018-08-20 | 2020-02-28 | 北京大学 | 利用抑制性tRNA系统制备PTC稳定细胞系及应用 |
WO2020181837A1 (en) * | 2019-03-13 | 2020-09-17 | Zhejiang Senwei Biopharmaceutical Development Co., Ltd. | Method for rescuing influenza virus and composition therefor |
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CN110743007B (zh) * | 2019-09-10 | 2023-05-30 | 安徽智飞龙科马生物制药有限公司 | 一种联合疫苗及其制备方法、应用 |
CN116904403A (zh) * | 2023-09-13 | 2023-10-20 | 北京民海生物科技有限公司 | 一种腮腺炎病毒突变体及其应用 |
CN116904403B (zh) * | 2023-09-13 | 2024-01-09 | 北京民海生物科技有限公司 | 一种腮腺炎病毒突变体及其应用 |
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CO2018006706A2 (es) | 2018-07-10 |
US11510974B2 (en) | 2022-11-29 |
JP2019500051A (ja) | 2019-01-10 |
CA3010015A1 (en) | 2017-07-06 |
BR112018013168A2 (zh) | 2018-12-11 |
CN106929482A (zh) | 2017-07-07 |
CN109563492A (zh) | 2019-04-02 |
US20200268870A1 (en) | 2020-08-27 |
EP3399029A1 (en) | 2018-11-07 |
CN109563492B (zh) | 2023-05-12 |
EP3399029A4 (en) | 2019-07-17 |
AU2016383201A1 (en) | 2018-07-05 |
ZA201804362B (en) | 2019-04-24 |
IL260258A (en) | 2018-07-31 |
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