Application of 6 Small Molecule Drugs in Inhibiting Canine Parvovirus

A technology of canine parvovirus and medicine, applied in the field of known compounds, can solve the problems of vaccine weakening

Active Publication Date: 2022-03-01
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although inactivated or attenuated live vaccines have been widely used for prevention, the repeated emergence of mutant strains has raised concerns and concerns about the effectiveness of existing vaccines, while maternal antibodies have also been shown to weaken vaccines

Method used

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  • Application of 6 Small Molecule Drugs in Inhibiting Canine Parvovirus
  • Application of 6 Small Molecule Drugs in Inhibiting Canine Parvovirus
  • Application of 6 Small Molecule Drugs in Inhibiting Canine Parvovirus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1. Determination of the protective effect of 6 small molecule drugs on CPV inhibition in F81 cells

[0062] Before F81 cells were purchased and used, 4 μL of small molecule drugs (Closantel / Closantel Sodium / Gemcitabine HCl / Trifuridine / Gemcitabine / Cladribine) (10 mM) and 4 μL of control drug Cidofovir (10 mM) were added to 156 μL of maintenance medium (MM), respectively, A 250 μM drug stock solution was prepared.

[0063]Drug treatment group: each drug stock solution was used to treat the F81 cells respectively. For each treatment group, 4 μL of 250 μM drug stock solution was added to 86 μL of F81 cells (25,000 cells per well), and the final concentration of each drug was 10 μM. After continuous treatment for 1 hour, 10 μL of CPV (New CPV-2a strain SD6) The drug-treated F81 cells were infected with an MOI (multiplicity of infection) of 0.076. Cell viability was assayed 40 hours after infection.

[0064] Positive control: DMSO was added to F81 cells to a final ...

Embodiment 2

[0069] Example 2. Determination of 50% effect concentration (50% antiviral efficacy, EC50) and 50% cytotoxicity concentration (50% cytotoxicity concentrations, CC50) of 6 kinds of small molecule drugs

[0070] The EC50 and CC50 of the drugs were determined by dose-response assay. details as follows:

[0071] The EC50 determination procedure is as follows: 86 μL of F81 cells (25,000 cells per well) were pretreated with 4 μL of doubly diluted drug (Closantel / Closantel Sodium / Gemcitabine HCl / Trifuridine / Gemcitabine / Cladribine) (final concentration range 0.3125-20 μM) for 1 h , and then 10 μL of CPV (New CPV-2a strain SD6) (MOI=0.076) was used to infect the drug-treated cells.

[0072] The CC50 determination procedure is as follows: 96 μL of F81 cells (25,000 cells per well) are mixed with 4 μL of doubly diluted drug (final concentration range is 0.3125-80 μM).

[0073] Both EC50 and CC50 determinations were performed in triplicate. After 40 hours of incubation, the Cell Coun...

Embodiment 3

[0077] Example 3. Immunofluorescence assay (IFA) detection of inhibition of viral VP2 protein expression by six small molecule drugs

[0078] Drug treatment group: 86 μL of F81 cells in a 96-well plate (25,000 cells per well) were mixed with 4 μL of doubly diluted drugs (Closantel / Closantel Sodium / Gemcitabine HCl / Trifluridine / Gemcitabine / Cladribine) (the final concentration of the drug was 5 μM, respectively, 10 μM and 20 μM) were pretreated for 1 hour, and the treated cells were infected with 10 μL CPV (New CPV-2a strain SD6) (MOI=0.076). Immunofluorescence assays were performed approximately 30 hours after infection.

[0079] Control group: DMSO was added to F81 cells in a 96-well plate to a final concentration of 0.1% (volume fraction), and an immunofluorescence test was performed 30 hours later.

[0080] Immunofluorescence test:

[0081] Fixation: Discard the culture medium in the 96-well plate and wash with 1×PBS (Gibco TM , USA, article number: 20012050) wash 3 times,...

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Abstract

The present invention relates to the new application of known compounds, especially the application of six small molecular drugs in inhibiting canine parvovirus. The invention provides the purposes of Closantel, Closantel Sodium, Gemcitabine HCl, Trifluridine, Gemcitabine and Cladribine in the preparation of the medicine for inhibiting the replication of canine parvovirus, the active ingredient of the medicine comprises Closantel, Closantel Sodium, Gemcitabine HCl, Trifluridine, Gemcitabine and / or Cladribine. The data prove that these 6 small molecule drugs all have excellent inhibitory effects on CPV replication, and can inhibit the expression of VP2 protein of different genotypes of CPV virus.

Description

technical field [0001] The present invention relates to new uses of known compounds, especially the use of Closantel, Closantel Sodium, Gemcitabine HCl, Trifluridine, Gemcitabine and Cladribine in inhibiting canine parvovirus. Background technique [0002] Canine parvovirus (Canine parvovirus, CPV) is a kind of single-stranded DNA virus with simple structure, which is a member of Parvoviridae. It has no envelope and forms an icosahedron. The total length of the viral genome is about 5300nt, containing two ORFs, the 5' end mainly encodes the regulatory proteins of early transcription (NS1 and NS2), and the 3' end encodes the structural proteins of late transcription, namely the viral capsid proteins (VP1 and VP2). The VP2 protein is the main component of the capsid protein, which can bind to the transferrin receptor (TfR) on the host cell membrane, thereby mediating the infection of the parvovirus. CPV is one of the main pathogens of canine acute gastroenteritis, leukopenia ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K31/7068A61K31/277A61K31/7076A61K31/7072A61P31/20
CPCA61K31/7068A61K31/277A61K31/7076A61K31/7072A61P31/20A61K2300/00Y02A50/30
Inventor 杨兵周宏专苏霞徐福洲林路路张进齐颀
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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