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MRNA and novel coronavirus mRNA vaccine containing same

A virus and -UTR technology, applied in the direction of DNA / RNA vaccination, positive single-stranded RNA virus, virus, etc., can solve the problems of poor curative effect, low expression efficiency, weak immune effect, etc., and achieve the benefit of large-scale industrial Production, high protein expression efficiency, and reduced immunogenicity

Pending Publication Date: 2022-05-13
SHENZHEN GINO BIOTECHNOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The main defect that inactivated vaccine exists at present is: 1) general immune effect is weak, can only induce humoral immunity; Different degrees of influence; 4) The vaccine response imbalance among the various antigenic components may induce other diseases
The main defects of adenovirus vector vaccines are: 1) the duration of the induced immune response is short; 2) lack of targeting, it may infect normal cells and cause adverse reactions; 3) if the body itself has immunity to adenovirus vector The adenovirus antibodies in the body will turn to attack the vector and make the vaccine ineffective
The main shortcomings of DNA vaccines are: 1) the expression efficiency may not be high; 2) the risk of viral gene integration into the host chromosome; 3) DNA needs to enter the nucleus, and then go through transcription, translation and other processes, the process is long and slow
[0008] Considering the above-mentioned traditional vaccines with low safety, poor curative effect, long production cycle and long time required to produce immunity, and the long time required for vaccine research and development, various vaccines have their own advantages, disadvantages and technical barriers. The safety and effectiveness of the vaccine have not yet been proven, and there is still a risk of failure. At present, there is still no new coronavirus pneumonia vaccine approved for marketing, and it has not been determined which vaccine has the best effect. Therefore, the development of new coronavirus treatment drugs and Vaccine, active prevention and treatment of novel coronavirus infection (SARS-CoV-2) is imminent

Method used

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  • MRNA and novel coronavirus mRNA vaccine containing same
  • MRNA and novel coronavirus mRNA vaccine containing same
  • MRNA and novel coronavirus mRNA vaccine containing same

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0090] ORF sequence optimization of embodiment 1 novel coronavirus SARS-CoV-2 mRNA

[0091] In this example, the whole genome data of 69 strains were collected from four public data sources (CNGBdb / GenBank / Genome Warehouse / GISAID), among which the data of 10 strains from GenBank can obtain their protein sequences at the same time, and the remaining 59 strains only have the whole genome For the data, the genome sequence was annotated using the virus genome ORF reader VIGOR, and the S protein sequences of the remaining 59 strains were obtained. VIGOR (Viral Genome ORF Reader) is a web-applied tool for gene prediction in influenza, rotavirus, rhinovirus, and coronavirus subtypes. VIGOR detects protein coding regions based on sequence similarity searches, and can accurately detect genome-specific features such as frameshifts, overlapping genes, embedded genes, and can predict mature peptides within a single polypeptide open reading frame. The program has built-in genotyping capab...

Embodiment 2

[0095] The 3'UTR sequence optimization of embodiment 2 novel coronavirus SARS-CoV-2mRNA

[0096] The 3'UTR sequence optimization scheme of SARS-CoV-2mRNA in this embodiment is shown in Table 1, specifically as follows:

[0097] The key element of the SARS-CoV-2 mRNA sequence is the conventional two-enzyme method Cap1 in the field (using the vaccinia virus capping enzyme and other components to add the 7-methylguanosine cap structure (Cap 0) to the 5' end of the RNA , and then use 2'-O-methyltransferase and the methyl group on SAM to transfer to Cap0 to form Cap1), the partial 5'UTR sequence of human ribosomal protein L32 (PRL32) (sequence shown in SEQ ID NO:3 : 5'-GGGGCGCTGCCTACGGAGGTGGCAGCCATCTCCTTCTCGGCATCAAGCTTACC-3'), ORF is the gene sequence encoding S1 protein (as shown in SEQ ID NO: 1) and polyA sequence (120 A, added in advance on the plasmid tail). The difference is that SARS-CoV-2-A mRNA adopts the 3'UTR sequence of human beta globin (HBB) (the sequence is shown in...

Embodiment 3

[0102] The optimization of the Poly (A) of embodiment 3 novel coronavirus SARS-CoV-2 mRNA

[0103] The sequence optimization scheme of the SARS-CoV-2mRNA in the present embodiment is as shown in Table 2, specifically as follows:

[0104] The key elements of the SARS-CoV-2 mRNA sequence are the partial 5'UTR sequence of Cap1 and human ribosomal protein L32 (PRL32) using the dual-enzyme method, and the ORF is the gene sequence encoding the S1 protein in the genome sequence of the strain whose GenBank is MN908947 (such as shown in SEQ ID NO: 1), human beta globin (HBB) 3'UTR sequence.

[0105] The sequence part of SARS-CoV-2-C and SARS-CoV-2-D is the same, the difference is that the SARS-CoV-2-C mRNA adopts the method of adding tail 120A in advance on the pcDNA 3.1(+) plasmid, SARS-CoV -2-D mRNA adopts the way of in vitro enzymatic tailing 120A. SARS-CoV-2-C is the same as SARS-CoV-2-E and SARS-CoV-2-F except for polyA, and also adopts the tailing on the plasmid in advance, the...

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Abstract

The invention discloses a separated mRNA (messenger Ribonucleic Acid), which comprises an mRNA for coding a new coronavirus S1 protein, and the amino acid sequence of the S1 protein is as shown in SEQ ID NO: 2; the invention relates to a polyadenylic acid derivative which is characterized by further comprising one or more of the following (a)-(d): (a) a 5 '-cap structure, (b) 3'-polyadenylic acid, (c) 5 '-UTR and (d) 3'-UTR, also disclosed are DNA, compositions comprising the same, liposome nanoparticles, mRNA vaccines against new coronavirus, pharmaceutical compositions, and kits. The mRNA can be highly expressed in cells; the method is economical, efficient, safer and efficient, the structure is more stable, the protein expression efficiency is higher, and the new coronavirus S1 protein can be continuously expressed. When the liposome nano-particles / vaccines are prepared from the liposome nano-particles / vaccines, an enough protection effect can be achieved by using an extremely small dosage, the immunogenicity is reduced, and meanwhile, an organism immune system can be activated to generate humoral immunity and cellular immunity.

Description

technical field [0001] The present invention relates to an mRNA, a DNA capable of transcribing the same, a composition containing the same, a liposome nanoparticle, an mRNA vaccine against a novel coronavirus, a pharmaceutical composition and a kit. Background technique [0002] New Coronary Pneumonia refers to pneumonia caused by a new type of coronavirus (SARS-CoV-2, referred to as New Coronary Virus). It is an acute infectious pneumonia. It has the ability to infect humans. In the early stage of infection, patients have symptoms of fever, fatigue, and dry cough. , In severe cases, dyspnea, respiratory distress syndrome or septic shock may occur. Coronavirus (Coronavirus) is a common and ancient virus family. There are 6 kinds of Human Coronavirus (HCoVs) confirmed today, including the SARS coronavirus (SARS) that causes acute respiratory syndrome (SARS). -CoV), MERS coronavirus (MERS-CoV) causing Middle East Respiratory Syndrome (MERS), and novel coronavirus (SARS-CoV-2)...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/50C12N15/85C12N5/10C07K14/165A61K39/215A61K39/39A61K47/69A61K47/60A61P31/14
CPCC07K14/005C12N15/85A61K39/12A61K39/39A61K47/6911A61K47/60A61P31/14C12N2770/20022C12N2770/20034A61K2039/53
Inventor 黄英李波汤勇杨锦霞李涛李冬丽张乐李佑平熊恒
Owner SHENZHEN GINO BIOTECHNOLOGY CO LTD
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