Classifications

• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K31/00—Medicinal preparations containing organic active ingredients
  • A61K31/33—Heterocyclic compounds
  • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
  • A61K31/53—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with three nitrogens as the only ring hetero atoms, e.g. chlorazanil, melamine
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/12—Antivirals
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
  • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
  • A61P31/12—Antivirals
  • A61P31/14—Antivirals for RNA viruses
  • A61P31/16—Antivirals for RNA viruses for influenza or rhinoviruses

Definitions

• the invention reiates to biologically active compounds possessing antiviral properties for the treatment and prevention of viral infections of animals and humans.
• the invention can be used in hospitals, research laboratories, as well as livestock and poultry.
• the closest in structure to the claimed compound from this series (1 ) can be regarded as a prototype.
• the compound (1 ) in a concentration of 40 ug / mi in in vitro experiments the infectious titer of influenza ⁇ / ⁇ 3 ⁇ 2 and influenza virus
• CC50 concentration at which 50% of the cells die
• the invention is the small molecule (2) which has antiviral activity.
• the small molecule (2) has demonstrated low toxicity.
• Fig. 1 shows a scaffold for a small molecule.
• Fig. 2 shows a small molecule based on the scaffold of Fig. 1 .
• the compound produced from this synthesis is a pale yellow crystaliine solid, soluble in water, methanol, dimethyl sulfoxide, insoluble in benzene, ether and most other solvents.
• the compound is orally ingestibie and show efficacy against viruses including the influenza virus.
• Viruses Used a one-day monolayer culture of epithelial cells MDCK (dog kidney) Viruses. To assess the antiviral activity of the virus used the reference A / Puerto Rico/8/34, as well as pandemic influenza virus H1 1 v A/San kt-Peterburg/2/09 (similar to the so-called virus "swine fiu» A/Caiifornia/7/09).
• the maximum tolerable concentrations of the compounds was determined by MTT test in ceil culture MDCK.
• Testing of toxicity was carried out as follows: weighed weigh weighing 5 mg in a sterile test tube 5 ml. and the diluted growth medium for the cells MDCK (a-MEM, Bioiot, St. Russia) to a concentration of 1 mg / mL, thus obtaining a basic solution. More of the same medium made eight consecutive binary dilutions (500, 250, 125, 82.5, 31 .25, 1 5.13, 7.56 and 3.78 mg / mi, respectively), which was used for toxicity testing. The experience set in the four parallels for each concentration. One-day cell culture MDCK, grown in 98 ⁇ vveli plates (Costar), checked visually in an inverted microscope on the integrity of the monolayer.
• the plates were washed twice with medium without serum, and then have made the test compound in appropriate concentrations in a volume of 100 ml in each well.
• the plates were incubated for 72 h at 37 ° C in the presence of 5% C02, and then recorded the results of the experiment visually assessing the integrity of the monolayer compared with control cells, and by the MTT (quantitatively evaluating the viability of the ceils) using the tablet reader Hydex Chameleon.
• Statistical analysis was performed using the program Statistica 6.0.
• the plates were incubated for 60 min at 37 ° C in the presence of 5% CO 2, and then washed with medium to remove unbound viral particles to cells. Continue to make the drug in the wells with virus dilutions in 100 mi of the appropriate concentration. Each concentration of test compound was placed in four parallels for each virus dilution. Control wells filled with growth medium in the same volume. Also, the wells were left to re-test toxicity used concentrations.
• the plates were incubated for 72 h at 37 0 G, and then recorded the results of the experiment visually assessing the integrity of the monolayer as compared to control ceils and the degree of cytopathic effect of virus in ceil culture, put hemagglutination reaction and MTT method was used to quantify cell viability using the tablet reader Hydex Chameleon.
• Evaluation of toxicity, as well as evaluation of the antiviral action of drugs produced by three methods 1 ) monitoring and evaluation of the integrity of the monolayer cells under an inverted microscope. Implies a comparison of cell morphology of control wells with experienced and registration of changes under the influence of an agent (drug, virus, etc.). The change of morphology could include violating the integrity of the monolayer, changes in cell shape, expression of cytopathic effect in virus infection.
• MTT The method used in the evaluation of drug sensitivity, based on the ability of dehydrogenases of living cells to restore the form of colorless 3- 4,5-dimethylthiazol-2-yl-2 ,5-difenilterarazola (MTT reagent) to blue crystalline farmazana soluble in DMSO or 96% alcohol. Saturation of color indicates the intensity of metabolic processes in cells, ie, the normal level of viability. The color intensity is recorded with a spectrophotometer and the absorbance values obtained can be used for statistical data processing.
• the concentration of the drug during that kills 50% of a monolayer of cells in a test on the MDCK cells was 177 pg / ml.
• Drug concentration at which all the cells survive the monolayer was 62.5 micrograms / ml, which was used to test the antiviral activity of the drug.
• connection (2) in much smaller concentrations, in comparison with toxic, has a pronounced antiviral activity, reducing the titers of virus A / Puerto Rico/8/34 virus and pandemic influenza HlNlv A/Sankt- Peterburg/2/09 (A/California/7/09-podobny) for 3,0 lg concentrations 62.5 and 125 ug / ml, respectively, then there is a more active antiviral compound than with a prototype.

Landscapes

• Health & Medical Sciences (AREA)
• Life Sciences & Earth Sciences (AREA)
• Virology (AREA)
• Chemical & Material Sciences (AREA)
• Medicinal Chemistry (AREA)
• Pharmacology & Pharmacy (AREA)
• Animal Behavior & Ethology (AREA)
• General Health & Medical Sciences (AREA)
• Public Health (AREA)
• Veterinary Medicine (AREA)
• Chemical Kinetics & Catalysis (AREA)
• Communicable Diseases (AREA)
• Oncology (AREA)
• General Chemical & Material Sciences (AREA)
• Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
• Organic Chemistry (AREA)
• Pulmonology (AREA)
• Molecular Biology (AREA)
• Epidemiology (AREA)
• Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
• Nitrogen Condensed Heterocyclic Rings (AREA)

Priority Applications (3)

Applications Claiming Priority (1)

Publications (2)

Family

ID=48984871

Family Applications (1)

Country Status (3)

Cited By (2)

Family Cites Families (8)

• 2012
  • 2012-02-14 JP JP2014557606A patent/JP2015507007A/ja active Pending
  • 2012-02-14 EP EP12868920.5A patent/EP2819517A2/en not_active Withdrawn
  • 2012-02-14 WO PCT/US2012/025070 patent/WO2013122575A2/en not_active Ceased

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