EP2437766A1

EP2437766A1 - Water-soluble antiviral product containing aframomum melegueta, for the treatment and prevention of acquired immunodeficiency syndrome (aids)

Info

Publication number: EP2437766A1
Application number: EP09786352A
Authority: EP
Inventors: Erick Vidjin' Agnih Gbodossou
Original assignee: Individual
Current assignee: Individual
Priority date: 2009-03-31
Filing date: 2009-03-31
Publication date: 2012-04-11
Also published as: CN102131513A; WO2010112966A1; US20120052140A1

Abstract

The invention relates to a water-soluble, virostatic, virolytic antiviral product containing Aframomum Melegueta, for the treatment and prevention of acquired immunodeficiency syndrome (AIDS) and the variants thereof. The product is produced as follows: once picked, the leaf of the Aframomum Melegueta is dried naturally and reduced to a fine powder; the dried powder is macerated in water sterilised with 100% methanol; and the solution is then passed through a filter of 45 microns in diameter, providing a water-soluble, virostatic, virolytic, sterilised antiviral product. Different experiments have shown that the product: has an antiviral activity; has a destructive toxic effect on cells infected by the AIDS virus; inactivates the virus and appears virostatic; is soluble in water; and is not complex and contains molecules smaller than 10,000 Da and is resistant to treatment of the protease.

Description

WATER-SOLUBLE ANTIVIRAL PRODUCT BASED ON AFRAMOMUM MELEGUETA FOR THE TREATMENT AND PREVENTION OF ACQUIRED IMMUNO DEFICIENCY SYNDROME (AIDS)

5 DESCRIPTION

The invention relates to an antiviral product based on Aframomum Melegueta virolysis, viro-static and water-soluble leaf. It is intended for the treatment and prevention of Acquired Immune Deficiency Syndrome (AIDS) and its variants. AIDS

10 is an infectious disease that strikes hard all over the planet and particularly African countries. It has been found that AIDS is often accompanied by lymphadenopathic syndromes which in their final stage are manifested by a collapse of the subject's immune defenses. The agent responsible for this pandemic has been isolated since 1983 by Professor Luc MONTAGNIER of the Institut Pasteur.

The prior art known to the inventor made it possible to identify the following documents:

OAPI Patent No. 09043 issued to CRUZ Joseph on March 31, 1991 and relating to new antiviral compositions based on extract of the Mimosa Pudica LINN leaf. OAPI Patent No. 11602, issued to Dr. Erick GBODOSSOU and relating to a composition of extracts

20 of 5 (five) plants (Momordica Balsamina, Aframomum Melegueta, Cyperus Articulatus, Ficus Iteophylla, Tamarindus Indica). In its therapy, the absorbed product, eradicates the AIDS virus through stool and urine. WO 94/18993 issued September 1994 to PHARMAKON USA, Inc. US Patent 5484889 issued University Apr. 16992. The patent WO 00 35 466 issued to (PAYA BIOTECHNIC INC) on June 22, 2000. The patent

US 5886029 issued to (KIRPAL S. DHALIWAL) March 23, 1999. All current research on the AIDS virus has not yet been completed either in vaccine development or in killing the virus. inside the body. Current antiretrovirals are mutating the pathogen into more and more virulent viruses. For more than twenty years, there has been a pandemic, research institutes,

30 Governmental and non-governmental agencies, United Nations Agencies, in short, the entire Scientific, Political, Cultural and Civil Society Community has invested in the fight against HIV / AIDS to find either a vaccine or an antiviral, often with the World Health Organization or other partners and donors such as the Ford Foundation. The conventional approach to treating HIV infection through the use of antiviral drugs such as azidothymide (AZT), DDC, 3TC, TAZT / 3C, indinavir reduces plasma viral load at the lower and as long as possible. The results, although positive, remain limited. On average 50% of patients have their plasma viral load dropped to 50 copies / ml after approximately 12 months of treatment. But in the current state of research, problems still unresolved by the use of antiretrovirals are still relevant, including the impossibility of total eradication of the pathogen. Immune recovery is insignificant even when the CD4 cell count has increased significantly, the permanence of the antiretroviral toxicity is not yet resolved.

Other problems are not yet solved by the use of antiretrovirals such as the risk of early development of antiretroviral resistance, the uncertainty as to the duration of maintenance of treatment efficacy. It should also be noted the constraints related to the use of current antiretrovirals, because it requires patients to absorb twenty products every day and their cost is very high compared to the purchasing power of African populations who are most exposed to HIV / AIDS infection. Furthermore, in the current state of the art, there is still no injectable, water-soluble antiretroviral product. No product of this type is known to date. One of the recommendations for prevention is the use of condoms, but we know that its use not only limits procreation but above all its effectiveness is not total. Increasingly, the international community is becoming aware of the need to involve healers in HIV prevention and to call on Traditional Medicine. The product, object of the invention, is based on leaves of Aframomum Melegueta, water soluble, virolysis and viro static. On the selected plant, studies on the botany, pharmacological and chemical aspects were carried out by the laboratory of the Ministry of Scientific Research of Senegal, and analyzes of the extract of the plant were made by the laboratory of Morehouse School of Medicine (Atlanta) USA, based on a repository developed by the inventor.

I - BOTANICAL, PHARMACOLOGICAL AND CHEMICAL STUDIES Name of the plant: Aframomum Melegueta.

Botanical aspect Rhizomatous plants with erect stems, up to 1 to 1.50 meters high. Beautiful flower in spikes located on a peduncle coming out of ground; Red fruits.

Chemical appearance Distillation produces essential oils containing Caryophyllene, Kayene, Pinene, Cineol, Geraniol, Geranyl Acetate and Terpineol. The seeds called maniguette contain 0.30% of a yellowish essence with pleasant odor, aromatic and a very bitter bitter principle: the neighbor Paradol of Gingerol. Its action polarizes the action of momorcharin by increasing the evolution of the inhibiting medium and neutralizing the viral proliferation.

Pharmacology aspect! eu

Aframomum are tenicidal purgatives and the decoction is used for baths in cases of deficiency edema. They have deworming properties from the roots. They play a big role in the treatment of rheumatism and lung diseases.

II - IN VITRO STUDY OF AFRAMOMUM MELEGUETA

The dried, powdered leaf was donated to the Virology Services team at Morehouse School of Medicine (Atlanta - USA) to verify its in vitro activity on the virus and its variants, based on the inventor's repositories. .

A - Objectives

It is a question of determining, on the one hand, whether the extracts of Aframomum Melegueta block the replication of the AIDS virus and, on the other hand, whether the dried powder of Aframomum Melegueta can be water-soluble. The research also aims to study the direct effect of the product on cells infected with Acquired Immune Deficiency Syndrome (AIDS) virus and its variants.

B - Implementation of tests

The purified fractions of the leaf extracts mixed in 1 mg of p24 equivalent to the virus (NL4-3KFS). The virus and the extract are tested initially from 10 mg per 1 microgram of extract. The extracts must be tested for a current application and show a direct effect on the virus. This could include blocking the protein from the envelope of the virus gρl20 or the dissolution of the lipids contained in the envelope of the virus.

Alternatively, it is conceivable that active ingredients can integrate virions and affect viral capsids or catalytic proteins such as reverse transcription, protease or integrase.

After a preliminary incubation of 15 minutes with the extract, the extract / virus mixture must be added to cultures of the tissue from which the support was taken. The virus must absorb it for 30 minutes at a temperature of 37 ° C. The culture is then washed 3 times with PBS and finally stored in fresh medium. For initial studies, the MAGI indicator cells that are HeLa cells that are designed to be infected with HIV-I should be used. Following infection, these cells produce beta-galactosidase, detectable as a blue color on the development of the infection. A comparison of the number of blue cells treated with untreated cells provides a relative indication of inhibition. The virus used in the initial studies must not have any gene and is therefore limited to a single stage of infection of the cell. This single stage of cell infection is by far the most appropriate approach for determining the effects of infection. It is important to establish in these tests an appropriate dose that is not toxic to the cells and that still contains a maximum concentration of soluble substance. If antiretroviral activity is found in the initial test, it will be necessary to determine how much of the viral life cycle is being affected. Specific viral isolators and "indicator" cells can be used to determine the relative performance of virus entry. If the entry of the virus is intact, it is possible to determine the extent to which reverse transcription has been affected by using PCR assays that detect different steps of viral DNA formation. The ability to produce intact virions can be directly determined using standard p24 ELISA assays while viral processing can be determined by viral protein pates present in the virions. It must also be able to at least make an initial determination of the nature of the soluble substance. Extrease protease treatment is used to determine whether the proteins present in the extract are responsible for the activity.

Based on the objectives and guidelines, experiments were conducted. III - EXPERIMENTS ON THE CHARACTERIZATION OF THE PRODUCT A- Solubility of the extract of Aframomum Melegueta

1 gram of dried Aframomum Melegueta powder is macerated in distilled water sterilized with 100% methanol and allowed to stand for one hour at room temperature. The resulting solution is then passed through a sterilization apparatus having a 0.45 micron filter to retain the remaining insoluble material.

The soluble fraction is devoid of bacteria. The substances present at the beginning in the dried materials are completely dissolved. The solution obtained is a maximum concentration of the dried powder of about 100 mg / ml.

B - Effect of the Product on infected cells

In order to determine whether the soluble extract has any antiviral activity, we tested its effect using the classic MAGI cell on infected cells.

MAGI cells are genetically engineered by HeLa cells that contain a reporter gene cassette comprising the viral LTR, located upstream of the E. coli • - gal encoding gene. - The expression of the gene is dependent on the activation by the HIV protein Tat, which requires an infection with HIV.

The infected cells turn blue and can be counted using a microscope.

- We inoculated 2 plates with 1 ng of HIV virus KFS. At the same time 200 μl of extract are added to the cells and placed in incubation for 2 hours.

The cells are washed twice and the extract is returned to fresh medium. The cells infected for 48 hours disappear in the presence of the extract of the product.

Given this, we can say that the extract acts directly on the virus and its effect is predominant on the infected target cells. To do this, we proceeded to the following test.

We made 2 other plates containing uninfected living cells. One of the plates is witness. In the 2 ^nd plate is brought on antiviral product based on Aframomum Melegueta. 30 minutes later, the AIDS virus is injected into the same plate containing the MAGI test cells before being ironed at the incubation temperature. These living cells did not turn blue, ie they were not infected. This experiment allows us to conclude that: - The product has antiviral activity.

The product inactivates the virus.

The product destroys cells infected with the Immuno Syndrome Virus

Acquired Disability (AIDS) and its variants.

C - Spectrometry of the product mixed by SELDI Mass

The product after study, showed that the active ingredients are not complex and the molecules therein are less than 10,000 Da in size and display a resistance to the treatment of the protease.

IV- RESULTS

These different experiments show that the product: has an antiviral activity. - has toxic and destructive effects on cells infected with the AIDS virus. inactivates the virus and appears as viro static. - is water soluble.

The product is not complex and has a set of molecules smaller than 10,000 Da in size and displays resistance to protease treatment.

The antiviral product based on Aframomum Melegueta for the treatment and prevention of Acquired Immune Deficiency Syndrome (AIDS) and its variants, object of the present invention is carried out as follows.

The Aframomum Melegueta leaf after collection is dried naturally and is reduced to a fine powder. The dried powder is macerated in water sterilized with 100% methanol. The solution is then passed through a filter 45 microns in diameter and gives an antiviral, virolysis, viro static and water-soluble product. The product can be used as a vaginal microbicide but also in any other form. It is demonstrated after the in vitro tests that the solution obtained blocks the development of Acquired Immune Deficiency Syndrome (AIDS) virus and its variants and indirectly stimulates and increases the reticuloendothelial system, thereby increasing the patient's immune defense.

Claims

1- Antiviral product based on AFRAMOMUM MELEGUETA and its method of preparation for therapeutic use for the Treatment and Prevention of Acquired Immune Deficiency Syndrome (AIDS) and its Variants.

2- Product according to claim 1, characterized in that its manufacturing process is in 3 steps: a) the first is the removal of the dried Aframomum Melegueta leaf and reduced to a fine powder. b) the second is the maceration of the powder of the Aframomum Melegueta leaf in 100% methanol sterilized water. c) the third is the passage of the solution in a 0.45 micron filter.

3- Product according to claim 2, characterized in that the product is hydrosubstituted.

4- Product according to claim 2, characterized in that the product has antiviral activity.

5. Product according to claim 2, characterized in that the product has toxic and destructive effects on cells infected with the virus Acquired Immunodeficiency Syndrome (AIDS) and its variants.

6- Product according to claim 2, characterized in that the product inactivates the virus and appears as virostatic.

7- Product according to claim 2, characterized in that the product is not complex and has a set of molecules less than 10,000 Da in size and displays a resistance to the treatment of the protease.

8- Product according to claims 1 to 7, characterized in that the product stimulates the reticuloendothelial system.

9- Product according to claims 1 to 8, characterized in that the product can be used as a vaginal microbicide and on any other form. 10- Product according to the set of claims, characterized in that the concentration and purification of the extract of the Aframomum Melegueta leaf is achieved by conventional methods used in chemistry and biochemistry.