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Antiviral agent and method for treating viral infection

A technology of antiviral agent and virus infection, applied in antiviral agent, gene therapy, biochemical equipment and method, etc.

Inactive Publication Date: 2019-09-06
黄立民
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] However, there is still an unmet need for broad-spectrum antiviral agents that can effectively treat infections caused by a variety of viruses regardless of the highly mutative characteristics of viruses

Method used

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  • Antiviral agent and method for treating viral infection
  • Antiviral agent and method for treating viral infection
  • Antiviral agent and method for treating viral infection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0072] Example 1: Morpholino oligonucleotides regulate MRJ splicing

[0073] The antisense morpholino oligonucleotide (MoMRJ) having the sequence shown in SEQ ID NO.1 is complementary to the 5' splice site of intron 8, and is used to interfere with the splicing of the MRJ gene. The potency of this morpholino oligonucleotide was assessed using an in vitro splicing assay. The MRJ pre-mRNA contains exons 8 to 9 and an internal truncated intron ( Figure 1A , top row). This MRJ pre-mRNA is spliced ​​in HeLa nuclear extracts. MoMRJ inhibits splicing, while the negative control (MoC) of morpholino oligonucleotides has no such effect ( Figure 1A , lower row). This result indicates that MoMRJ specifically disrupts intron 8 splicing. Next, the effect of MoMRJ on the expression of MRJ isoforms in HEK293T cells was evaluated. RT-PCR and Western blot analysis showed that increasing the amount of MoMRJ inhibited the inclusion of exon 9 / 10, thereby reducing the expression of MRJ-L mRNA...

Embodiment 2

[0074] Example 2: Morpholino oligonucleotides targeting MRJ inhibit HIV-1 replication

[0075] MoMRJ blocks HIV-1 replication in macrophages by interfering with MRJ splicing and inhibiting MRJ-L expression. As observed in HEK293T cells, MoMRJ reduced MRJ-L mRNA and protein expression in THP-1 cells, while MoC had no such effect ( Figure 2A ). HIV-1 infected THP-1 derived macrophages (Konopka and Duzgunes, 2002) were treated with MoMRJ and the expression of HIV core protein p24 was assessed. Immunoadsorption assays showed that MoMRJ significantly reduced the expression of p24 in the supernatant of HIV-1 infected cells, while MoC had no such effect ( Figure 2B ). The effect of MoMRJ in the early stages of HIV-1 infection was further evaluated using the HIV single-infection system in which the VSV-G pseudotype HIV-1 NL4-3 strain contains a murine thermostable reporter gene located in the nef region Antigen CD24 (HSA) gene (He et al., 1995). HSA positive cells were assessed...

Embodiment 3

[0076] Example 3: Morpholino oligonucleotides targeting MRJ inhibit RSV replication

[0077] The ability of MoMRJ to limit RSV production was further examined. MoMRJ and control MoC were tested for potency and used in Hep2 cells. RT-PCR and immunoblotting analysis showed that MoMRJ effectively reduced the mRNA and protein expression of MRJ-L, but did not have this effect on MRJ-S ( Figure 3A ). Subsequently, the amount of RSV virus in the morpholino oligonucleotide-treated cells was assessed. Immunoblot analysis showed that, in cells treated with MoMRJ, RSV F protein expression decreased significantly ( Figure 3B ). Phage plaque assay and RT-qPCR of RSV N mRNA confirmed that MoMRJ substantially inhibited virion production ( Figure 3C ). When treated with MoMRJ, viral subgenome mRNA production also decreased, while MoC showed no inhibitory effect ( Figure 3D). Therefore, MoMRJ inhibits RSV subgenome mRNA production by reducing the mRNA and protein expression levels ...

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Abstract

The present invention relates to antiviral agents and methods of their use in suppression of viruses and in the treatment of a disease or condition associated with viral infection. The antiviral agentincludes a nucleotide derivative is a morpholino oligomer complementary to mammalian relative of DnaJ (MRJ) gene.

Description

[0001] [Cross Reference to Related Application] [0002] This application claims priority to US Provisional Application 62 / 449,600, filed January 24, 2017, which is hereby incorporated by reference in its entirety for all purposes. technical field [0003] The present invention relates to an antisense oligonucleotide for treating virus infection and an antiviral treatment method using the oligonucleotide. Background technique [0004] Bacterial and viral infections are major threats to human health (Morens and Fauci, 2013). The treatment of bacterial infections largely relies on antibiotics (Bassetti et al., 2016; Bush and Bradford, 2016), while antiviral therapy is still mainly supportive and symptomatic. In addition, due to the continuous development and civilization of undeveloped areas, emerging and re-emerging pathogenic bacteria continue to pose threats to humans. Due to the lack of ready-to-use antiviral agents, humans will not be able to cope with sudden epidemic v...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K31/712A61K45/06A61P31/14A61P31/20A61P31/18A61P31/16
CPCC12N15/113C12N2310/11C12N2310/3233C12N2320/33A61P31/12A61K31/712Y02A50/30A61P31/14A61P31/18A61K31/245A61K31/522A61K31/655A61K31/7072A61K38/21C12N15/1131C12N15/1132C12N2320/30C12N2320/31
Inventor 黄立民柯释涵
Owner 黄立民
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