BRPI0709452A2 - concentration of immunoglobulins and f (ab) '2 and / or fab fragments specific to an arboviruses as a medicinal product - Google Patents

Abstract

Concentrate of immunoglobulins and F (ab) '2 or Fab arbovirus-specific Fabs as a medicinal product The invention relates to a new medicinal product for the treatment of arboviruses, that is, a concentrate of immunoglobulins and F (ab) fragments '2 and / or Fab specific to said arbovirus, as well as its preparation process.

Description

Patent Descriptive Report for "Arbovirus-specific F (ab) '2 and / or Fab immunoglobulin concentrate and / or Fab fragments as a medicinal product"

The present invention relates to a novel medicinal product for the treatment of arbovirus, that is, an immunoglobulin concentrate and F (ab) '2 and / or Fab fragments specific to said arbovirus, as well as their preparation process.

Introduction

Viruses that use arthropod vectors in their cycle are grouped under the general term arbovirus. Arboviruses are defined by WHO as viruses that subsist in nature essentially or mainly through biological transmission between vertebrate guests susceptible to hematophagous arthropods; they multiply and cause vertebrate viremia, proliferate in arthropod tissues, and are transmitted to another vertebrate by the stinging insect after an extrinsic incubation period.

Transmission of the virus from a viremic guest to an adult female mosquito occurs through the blood that is sucked in when the bite occurs. The virus multiplies within the mosquito, crosses the animal's stomach barrier and is found in the salivary glands. The contamination of a healthy human is acquired by the mosquito's anticoagulant saliva, released just before the sting in a blood vessel. The window during which a person is a viremic guest before becoming ill is only a few days.

Known arboviruses belong to five virus families:

Togaviridae, genus Alphavirus, 28 viruses including Chikungunya, O'Nyong Nyong viruses, Ross River, Sindbis and Mayaro.Flaviviridae, genus Flavivirus, 68 viruses, including yellow fever, dengue, Japanese encephalitis, West Nile virus, Temperate Eurasian tick-borne encephalitis virus and viruses causing Kyasanur forest disease and Omsk.Bunyaviridae, Bunyavirus genus (138 viruses, including Bunyamwera virus), Phleboviris genus (43 viruses, including Rift Valley fever), genus Nairovirus (24 viruses, including Crimea-Congo haemorrhagic fever virus) + 41 unclassified viruses.

- Reoviridae1 genus Orbivirus (69 viruses) and genus Coltivirus (2virus) + 6 unclassified viruses

Rhabdoviridae, genus Vesiculovirus (18 viruses) and genus Lyssavirus (16 viruses) + 36 unclassified viruses.

The main arboviruses observed in the tropics are detailed below.

Dengue is spread throughout the tropical and subtropical areas of the world and represents the main public health problem posed by arboviruses. There are four viral serotypes, called "dengue 1, 2, 3 and 4", which do not produce cross protection. Clinically, several forms of dengue are distinguished: asymptomatic dengue, classical dengue (CD) and severe forms, particularly hemorrhagic forms, hemorrhagic dengue (HD) and dengue shock syndrome (DSS), which can cause death, especially in children. All 4 types of dengue virus can be responsible for CD as well as HD. The pathophysiological mechanisms involved in the genesis of HD are unknown. The most common theory refers to the phenomenon of "immune facilitation": Since an individual who has become infected with one of the four serotypes is not protected against the other three, a heterologous secondary infection can lead to HD. Vectors are mosquitoes of the genus Aedes: Aedes aegypti is the main vector and Aedes albopictus plays an important role in peripheral rural and urban areas, and is well adapted to temperate climates.

West Nile virus is currently considered the most widespread of flaviviruses after dengue virus; It affects humans sporadically or epidemically. It was recently described as first emerging on the American continent in an epidemic that occurred in New York in 1999 (62 cases including 7 deaths). It then spread considerably in the United States, affecting more than 9,000 people in 44 states in 2003, including 2,866 cases of encephalitis and 264 deaths. The virus was previously found in various regions of the world, in Africa, the Middle East, India and Europe. Mosquitoes are the main vectors of West Niio virus, especially those of the genus Culex. The main hosts of the virus are birds, wild as well as domestic (ducks, pigeons, etc.). They play a key role in the spread of the virus. West Nile virus mainly affects humans via a mosquito vector. Infection is characterized by the sudden onset of a high fever after 3 to 6 days of incubation. This fever is accompanied by headaches and back pain, muscle pain, cough, swollen lymph nodes in the neck and often rash, nausea, abdominal pain, diarrhea and respiratory symptoms. In less than 15% of cases, complications occur: meningitis, encephalitis, and rarely hepatitis, pancreatitis, or myocarditis. Usually, the patient recovers spontaneously, sometimes with sequelae. However, the disease can be fatal in the elderly and sometimes in young children.

Yellow fever (YF) remains a formidable epidemic and a constant threat in sub-Saharan Africa and tropical America. It is due to the yellow virus. It manifests in the form of a hemorrhage, amarylline typhus, with an early or "red" phase, remission on the third day and a stasis or "yellow" phase, with jaundice, vomiting, hemorrhage (mainly in the GI tract) and renal syndrome. The epidemiology is complex due to the fact that, in natural areas, the amaril virus circulates continuously among monkey populations due to simiophilous wild mosquitoes acting as vectors. It was only by accident, when humans came into contact with this jungle cycle, that the first human cases occurred.

Chikungunya (abbreviated as CHIK) is transmitted by Aedes mosquitoes. The name originates from the Bantu1 language and means: the bending, curling, or bent man's disease, because it causes very severe joint pain combined with stiffness, which gives infected patients a very characteristic bending appearance.

Among more than 950 mosquito species, several can transmit chikungunya, but only Aedes aegypt and Aedes albopictus have been identified to date as epidemic vectors due to their adaptation to human habitation areas. These same species are also involved in the transmission of other arboviruses: dengue fever, dengue hemorrhagic fever (HDF) 1 yellow fever, etc.

The clinical profile is dominated by a high fever similar to that of dengue fever (dengue is often mistaken for chikungunya and vice versa), combined with disabling joint pain and sometimes rash. However, there are severe forms that have been ignored to this day: fulminant hepatitis, heart attacks, meningoencephalitis, etc. Several other alphavirus arboviruses (approximately 30 kD capsid and 3 'polyadenylated RNA) such as Ross River, O'nyong-nyong and Mayaro have been associated with similar symptoms.

The incubation of the disease lasts four to seven days on average. Viremia, the presence of the virus in the blood and therefore of possible transmission extends for approximately five days. Antibodies then develop. These remain in the blood. Therefore, immunity is usually acquired for life.

A vaccine is available to prevent yellow fever. Vaccination is systematic in exposed populations. However, 60 to 80% of the population needs to be immunized (naturally or through vaccination) to prevent epidemics. Antibodies appear after about ten days. Vaccination is contraindicated in pregnant women and children under 6 months of age.

There is also a vaccine in use against tick-borne encephalitis in Europe. It is sold under the name TICOVAC® (Baxter SA).

A phase I and a phase II study were performed in the United States for a chikungunya vaccine by the United States Army Medical Research Institute of Infectious Diseases (Edelman R et al. "Phase II study of safety and immunogenicity of live chikungunya virus vaccine" TSI-GSD-218 (June 2000; Am J Trop Med Hyg, 62: 681-5).

There is currently no treatment with arbovirus viricide.

Treatment is purely symptomatic, to lower fever and reduce pain.

Several studies have demonstrated the efficacy of immunoglobulin injections containing anti-hepatitis A antibodies in individuals at risk for exposure to this virus prior to the development of the hepatitis A vaccine (Ohara et al., Jpn J Exp Med1 October 1986; 56 (5): 229-33; Conrad ME et al., J Infect Dis, July 1987; 156 (1): 56-63).

Hepatitis B-specific immunoglobulins, or anti-HBs, are widely used to protect any unvaccinated person who has been cut with a contaminated object, newborns of mothers who are HBs antigen positive (in which case, an injection should be given immediately and should be accompanied by initiation of vaccination), liver transplant patients to avoid transplant reinfection, and sexual partners of HBs antigen positive individuals while waiting for vaccination to take effect. These immunoglobulins allow protection to be established prior to risk exposure or within 24 hours of contact with infection (accidental puncture).

Objectives and brief description of the invention

Given this lack of established viricide treatment, and only one vaccine having received marketing authorization, the applicant sought to develop a medicinal product to treat or prevent arboviruses based on specific immunoglobulins that enable rapid immunization of exposed persons.

We have surprisingly demonstrated that such treatment requires the combination of immunovulins and arbovirus-specific F (ab) '2 and / or Fab fragments to be treated to be effective.

Definitions

The term "concentrate" refers to a product obtained by the elimination of certain components. An immunoglobulin concentrate is obtained by eliminating certain plasma components to obtain an immunoglobulin enriched plasma fraction. The term "immunoglobulin" (Ig) refers to a natural globulin, present mainly in plasma, with antibody functions, which can be used in curative or preventive therapy.

Immunoglobulins are heterodimers composed of 2 heavy chains and 2 light chains, disulfide bonded. Each strand is comprised at the N-terminal position of a variable domain or region (encoded by rearranged VJ genes for the light chain and VDJ for the heavy chain) which is specific for the antigen against which the antibody is directed and at the C-terminal position of a constant region composed of a single CL domain for the light chain or 3 domains (CH1, CH2 and CH3) for the heavy chain. The combination of light and heavy chain variable domains CH1 and CL domains form the Fab parts, which are connected to the Fc region by a very flexible hinge region that allows each Fab to bind to the target antigen while the Fc region, intermediate of the properties. of effector antibody remains accessible to effector molecules such as the FcyR and C1q receptors.

IgG's are the most common immunoglobulins (75 to 80% of antibodies in the circulation). They protect the body against bacteria, viruses and toxins circulating in the blood and lymph. In addition, they quickly bind to complement (one of the components of the immune system). They also participate in memory response, which is the basis of immunity upon which the vaccination mechanism is based. Finally, immunoglobulins G cross the placental barrier and thus produce passive immunity in the fetus.

IgA's are found mainly in secretions such as saliva, intestinal juice, sweat and breast milk. The primary role of immunoglobulins A is to prevent pathogens from binding to cells, particularly the protective cells that make up the mucous membranes and epidermis.

IgMs are immunoglobulins secreted at the body's first contact with an antigen. They are the first type of plasma cell-released immunoglobulins. The presence of IgM in the blood indicates an ongoing infection.

Enzymatic proteolysis of papain immunoglobulins generates 2 identical fragments, which are known as Fab fragment (Antigen-Binding Fragment) and an Fc fragment (crystallizable fraction). The Fc fragment supports the effector functions of immunoglobulins.

By pepsin proteolysis, an F (ab) '2 fragment is generated, in which the two Fab fragments remain linked by two disulfide bridges, and the Fc fragment is broken down into several peptides. The F (ab) '2 fragment is formed by two Fab' fragments (one Fab 'fragment consists of a Fab and a hinge region), linked by intercatenary disulfide bridges to form an F (ab)' 2.

The term "chromatography" refers to a method of separating the components of a mixture based on their selective retention using a suitable medium.

Detailed Description of the Invention

First, the invention relates to an immunovulin concentrate and arbovirus-specific F (ab) '2 and / or Fab fragments as a medicinal product.

The use of immunoglobulin enriched human plasma fractions for the treatment of various infections and congenital deficiencies has been known since the development of Cohn's ethanol precipitation process (Cohn et al. 1946, J. Am. Chem. Soc. 68, 459 Oncley et al., 1949, J. Am. Chem. Soc.71, 541).

Such F (ab) '2 or Fab fragments, which contain the antibody binding site, may have lost a number of the properties of the integral antibody from which they are derived, such as the ability to bind to Fcgam receptors.

The arbovirus in question, which may be treated with a concentrate as provided for in the invention, may be, for example, one of dengue viruses, yellow fever virus, West Nile virus, chikungunya virus, Ross River, O'Nyong-nyong virus or Mayaro virus.

In the concentrate according to the invention all combinations between a mixture of immunoglobulins A, G and / or M and F (ab) '2 and / or Fab fragments of arbovirus-specific Ig A, G and / or M are possible. Particularly, the concentrate according to the invention is a concentrate of immunoglobulins A, G and M and arbovirus-specific F (ab) '2 and / or Fab fragments.

Preferably, the concentrate according to the invention is an exclusively M-immunoglobulin concentrate, and of arbovirus-specific F (ab) '2 and / or Fab fragments.

Particularly preferably, the concentrate according to the invention is comprised of a concentrate exclusively of immunoglobulins G and F (ab) '2 and / or Fab fragments of arbovirus-specific IgG and IgM.

Preferably, the concentrate according to the invention contains at least 50% IgG immunoglobulins and 90 to 98% proteins that react with antibodies specifically directed against human immunoglobulins, particularly from 5 to 50% F (ab) '2 and / or Fab, particularly at least 50 to 60 g / l Ig and fragments for a pharmaceutical preparation.

According to the invention, from 1 to 10 mmol of magnesium and / or zinc may be added to the concentrate.

Another object of the invention is the use of a concentrate according to the invention for the manufacture of a medicinal product for the treatment of said arbovirus.

This treatment is prophylactic and / or curative. It is used to give passive immunity to people not yet infected in an epidemic region, or to treat patients already infected with the virus.

The medicinal product in question is administered topically, orally, mucosally, intramuscularly or intravenously.

It is effective for several weeks, approximately 21 days, after which time this administration should be repeated if the epidemic or symptoms persist.

The invention also relates to a process for preparing a concentrate according to the invention. This process comprises mixing an arbovirus-specific immunoglobulin concentrate and a proteolytic arbovirus-specific immunoglobulin concentrate to obtain F fragments. (ab) '2 and Fab specific for this arbovirus. This process therefore requires the preparation of at least one immunoglobulin concentrate.

This process begins by creating a pool of at least 1000 plasma donations, each donation containing a sufficient Ig titer directed against said arbovirus. A serum containing a sufficient titer corresponds, for example, to a serum that remains positive for detection of anti-chikungunya antibodies after dilution to 1/1000 when the titer is measured by a method such as Elisa.

These donations come from people who have been in contact with the disease, or patients who developed the disease.

Titration may be performed according to the procedure described by C. van de Water et al., Journal of Immunological Methods, 166 (1993), 157-164.

To enrich this plasma pool with immunoglobulins, the other plasma components known as "lipid and protein contaminants" are precipitated in one step. This one-step precipitation purification can be accomplished by plasma dilution under precipitation conditions according to Steinbuch (Steinbuch M., Archiv. Biochem. Biophys., 134, 279-284) and by the addition of caprylic acid. It can also be obtained by the addition of precipitating agents such as Rivanol, aluminum chloride, cetylpyridinium chloride, octanoic acid, polyphosphates and in the presence of adsorption agents such as tricalcium phosphate and bentonite.

The supernatant resulting from precipitation may constitute the immunoglobulin concentrate. It therefore contains a mixture of IgG, IgA and IgM. This supernatant is recovered, for example, by filtration, optionally with the addition of at least one filtering additive.

The supernatant resulting from centrifugation or filtration may undergo viral deactivation processing such as conventional solvent / detergent deactivation processing (Triton X100). If the precipitation performed was a "caprylic" precipitation such as As described above, caprylic acid residues in the supernatant are eliminated by calcium P04.

To obtain an IgG1 IgA or IgM1 concentrate the method described in patent application EP1385886 can be applied, particularly methods corresponding to pH adjustment, adsorption on a preloaded column, adsorption on the supernatant column containing immunoglobulins and proteins. accompanying them, column washing and sequential elution of the various immunoglobulin categories such as IgG, A or M. After the viral deactivation step, the supernatant then undergoes an additional chromatographic purification step on an anion exchanger performed. at alkaline pH. Particularly, the pH of the supernatant is adjusted above to a pH ranging from 8.9 to 9.1, and the column is loaded with a buffer at a pH of 8.9 to 9.1. The chromatography step allows adsorption of immunoglobulins on the column and passage of non-retained proteins into the effluent. Chromatography may be performed, for example, on a cross-linked polysaccharide or vinyl polymer gel grafted with DEAE, TMAE or QAE groups.

After washing the column with the same buffer as the loading buffer to remove non-retained proteins, immunoglobulins G are eluted with a phosphate buffer whose pH ranges from 4 to 7, preferably pH 6.2.

A subsequent optional elution with the same phosphate buffer supplemented with 100 to 175 mM NaCl, preferably 150 mM, at a pH of 6 to 6.3 can be used to collect IgA.

An optional subsequent elution with the same buffer adjusted to a pH ranging from 6 to 7 and supplemented with 250 to 350 mM NaCl, preferably 300 mM, can be used to collect IgM.

Any type of IgA1 IgG and IgM mixture can be considered by mixing the concentrates as described above.

Immunoglobulins thus eluted and collected can be concentrated by ultrafiltration and subjected, for example, to conventional sterilization filtration and then filtration through nanometric filters with porosity decreasing from 100 to 15 nanometers.

To the concentrated and filtered immunoglobulin solution is added a pharmaceutically acceptable stabilizing agent such as those described in patent application WO2004 / 091656, and then that solution is packaged as a sterile solution and optionally frozen and / or lyophilized.

The application of nanofiltration makes it possible to eliminate viruses resistant to solvent / detergent viral deactivation treatment.

A portion of the immunoglobulin concentrate thus obtained, or other immunoglobulin concentrate obtained as described above, is subjected to proteolysis to obtain F (ab) '2 or Fab fragments. The IG concentrate and the mixture of fragments resulting from proteolysis are then mixed.

Thus, an IgG and F (ab) '2 and / or IgG and IgM Fab concentrate are obtained.

(1) preparation of an IgG concentrate described above,

(2) preparation of an IgM concentrate described above,

(3) mixing and digesting a part of the IgG concentrate and a part of the IgM concentrate to obtain a mixture of IgG and IgM F (ab) '2 and / or Fab fragments,

(4) mixture of (1) and (3).

To obtain F (ab) '2 fragments, proteolysis is performed at pH 4.0 at 35 ° C with 1% pepsin, this percentage corresponding to the weight ratio of pepsin to the total protein weight of the concentrate (IGLOO protocol). ).

To obtain Fab fragments, 1% papain proteolysis is performed, this percentage corresponding to the ratio of papain weight to total protein weight of the concentrate.

Proteolysis of G, A and / or M immunoglobulins can also be performed using plasmin and / or trypsin, and the implementation of such proteases is well known to those skilled in the art. The example disclosed below describes a specific embodiment of the invention. but should not be considered as a limitation of its scope.

Example 1: Preparation of an anti-chikungunya immunoglobulin concentrate

1-1. Creating a plasma pool

One liter of anti-chikungunya antibody-rich plasma is collected from volunteer donors recently infected with the chikungunya virus and cured of disease symptoms. The antibody titer is measured by Elisa, and consists of fixing virus antigens on a microtiter plate, and then revealing the specific antibodies through a strong root peroxidase labeled reagent directed against immunoglobulins. For plasma pool creation, positive test samples are stored at a dilution of at least 1/1000 in the context of a "specific" Elisa method.

1-2. Preparation of immunoglobulins

The resulting plasma pool from step 1-1 is cooled to -3 ° C and during cooling, ethanol is added in a volume sufficient to achieve a final ethanol concentration of 8%. The precipitate that forms in it is discarded.

The pH of the supernatant is then adjusted to pH 5.9 by the addition of an acetate buffer, for example cooled to -5 ° C and supplemented with sufficient ethanol volume to obtain a final ethanol concentration of 19%. The precipitate formed therein is collected by centrifugation, for example, and resuspended in acetate buffer, for example, to obtain a final pH of 4.7 to 4.9.

Octanoic acid is then added at 20 ° C under intense stirring to obtain a final octanoic acid concentration of 20 g / l.

The precipitate formed therein is separated by centrifugation or alluvial filtration and discarded. Tricalcium phosphate or activated charcoal is added to the supernatant and then the mixture is clarified by deep bed filtration. The pH of the immunoglobulin-containing supernatant resulting from the clarification step is adjusted to 9 by the addition of NaOH / glycine buffer, for example, and the supernatant is applied to an anion exchange column (TMAE Fractogel, for example), which is loaded at pH 9 with a glycine / NaCl buffer at pH 9.

The loading buffer is flushed until the 280 nm column output OD [outer diameter] is close to the OD2so measured at baseline establishment.

IgG is then eluted with a first sodium phosphate buffer at pH 6.2. A second elution is performed with a phosphate buffer supplemented with 300 mM NaCl.

The corresponding eluate contains IgA, IgM and part of lgG4. The detailed operation process of such purification is disclosed in EP 1385886.

1-3. Preparation of an active concentrate against chikungunya

25% of the first eluate containing IgG1 is extracted and added to eluates containing IgG4, IgA and IgM. This immunoglobulin mixture is concentrated at 50 g / l by ultrafiltration into a membrane whose shear threshold is equal to or lower than 30 kD.

The pH of the concentrated mixture is adjusted by diafiltration against a citrate buffer at pH 3.8 to 4.2 to obtain an acidic pH within this range. The solution is then supplemented with pepsin (10,000 FlP / mg) so that the amount of pepsin corresponds to 1% of the total amount of protein contained in the concentrated mixture.

This solution is then filtered under sterile 0.2 mm conditions and incubated for 20 h at 37 ° C.

After incubation, the protein hydrolyzate is neutralized, for example, by the addition of sodium hydroxide at pH 6.2 +/- 0.2. The neutralized protein hydrolyzate is diafiltered against a glycine buffer at pH 6.2 +/- 0.2, until OD2SO reaches about 0.005, with OD28O being measured in the 30 kD cutoff membrane filtrate line. .Peptides resulting from pepsin proteolysis, the size of which is equal to or less than 30 kD, are discarded upon passage through the cut-off membrane. The protein hydrolyzate thus obtained contains Fab fragments, F (ab) '2 fragments but lacks Fc fragments.

The resulting protein hydrolyzate is then mixed with the remaining 75% of the first eluate containing IgG. The mixture is then concentrated by ultrafiltration to reach a final concentration ranging from 50 to 160 g / l, depending on the selected route of administration. The titer of the concentrate is measured according to the method described in Edelman, R et al. (American Journal of Tropical Medicine and Hygiene, 62 (6), 2000, pages 681-685). The titer thus obtained in anti-chikungunya specific antibodies of the concentrate is at least 3 to 10 times higher than that of the initial plasma.

1-4. Use of the preparation

The concentrate resulting from step 1-3 is stabilized by admixture with a formulation comprising pharmaceutically acceptable excipients, such as glycine at a final concentration of 0.22 M, or as described in patent application WO 2004 / 091 656. The pH of the formulation added to the concentrate is compatible with obtaining a liquid mixture whose pH ranges from 4.2 to 5.6.

The resulting liquid mixture may be administered, for example, intravenously, subcutaneously or intramuscularly, depending on the phlebological state of the recipient.

The administered dose is 0.2 to 0.8 ml / kg and may, in the event of an epidemic, be given as a precautionary measure every 3 weeks to patients who are particularly exposed, for example to the elderly, pregnant women or neonates.

Claims (23)

1. Immunoglobulin concentrate and arbovirus-specific F (ab) '2 and / or Fab fragments as a medicinal product. Concentrate according to Claim 1, characterized in that said arbovirus is one of the dengue viruses, yellow fever virus, West Nile virus, chikungunya virus, Ross River virus, O'nyong-nyong or the Mayaro virus. Concentrate according to Claim 1 or 2, characterized in that the immunoglobulins are immunoglobulins A, G and M. Concentrate according to Claim 1 or 2, characterized in that the immunoglobulins are G immunoglobulins. Concentrate according to claim 1 or 2, characterized in that the immunoglobulins are immunoglobulins M. Concentrate according to any one of Claims 1 to 5, characterized in that it contains from 90 to 98% immunoglobulin and F (ab) '2e / or Fab. Concentrate according to any one of claims 1 to 6, characterized in that it contains from 5 to 50% F (ab) '2 and / or Fab. Concentrate according to any one of claims 1 to 7, characterized in that the F (ab) '2 and / or Fab fragments are F (ab)' 2 and / or Fab fragments of IgG and / or IgM. Concentrate according to any one of claims 1 to 8, characterized in that from 1 to 10 mmol of magnesium is added thereto. Concentrate according to any one of claims 1 to 9, characterized in that from 1 to 10 mmol of zinc is added thereto. Use of a concentrate as defined in any one of claims 1 to 10 for the manufacture of a medicinal product for treating said arbovirus. Use according to claim 11 for the manufacture of a medicinal product in a form to be administered topically, subcutaneously, orally, intramuscularly or intravenously. A process for preparing a concentrate according to any one of claims 1 to 3, wherein said process comprises the following steps: - creating a pool of at least 1,000 plasma donations, each donation containing a sufficient Ig titer. directed against said arbovirus, - precipitation of lipid and protein contaminants in one step, - recovery of an Ig concentrate in the supernatant (1), - subjecting part of the first concentrate to proteolysis to obtain F (ab) '2 fragments and / or Fab (2), - mixture of fractions (1) and (2). Process for preparing a concentrate according to any one of claims 1 to 5, characterized in that it comprises the following steps: - creating a pool of at least 1,000 plasma donations, each donation containing a sufficient Ig titer. anti-chikungunya, - precipitation of lipid and protein contaminants in one step, - chromatography of the supernatant on an anion exchanger at alkaline pH, - elution of IgG with a phosphate buffer at pH 4 to 7 to obtain a concentrate IgG (1), - optionally then eluting IgA with the same 100 to 175 mM NaCl phosphate buffer added thereto, preferably 150 mM, at a pH of 6 to 6.3 (1), - optionally then eluting IgM with the same phosphate buffer at a pH between 6 and 7 and with 250 to 350 mM NaCl added thereto (1) .- optionally mixing IgG, IgA and IgM concentrates (1), - subjecting part of the first concentrate to proteolysis for obtain F (ab) '2 and / or Fab (2) fragments, - mixing fraction (1) with fraction (2). Process according to Claim 14, characterized in that the pH of the supernatant is adjusted to the range between 8.9 and 9.1 and the chromatography column is loaded with a buffer at pH 8.9 to 9. 1 before chromatography. Process for preparing a concentrate according to claim 4 and 8, characterized in that it comprises the following steps: (1) preparing an IgG concentrate according to claim 14 or 15, (2) preparing a IgM concentrate according to claim 14 or 15, (3) mixing and subjecting a portion of the IgG concentrate and an IgM concentrate part to proteolysis to obtain a mixture of F (ab) '2 and / or Fab fragments of IgG and IgM, (4) mixture of (1) and (3). Process according to any one of claims 13 to 16, characterized in that the proteolysis to obtain F (ab) '2 fragments occurs in 1% by weight pepsin of proteins at pH 4 and 35 ° C. Process according to any one of claims 13 to 17, characterized in that the proteolysis to obtain the Fab fragments takes place in papain. Process according to any one of claims 13 to 18, characterized in that the precipitation is a caprylic precipitation and in which the caprylic acid residues in the supernatant are removed by calcium P04. Process according to any one of claims 13 to 19, characterized in that the precipitate is filtered off after the addition of at least one filtering additive. Process according to any one of claims 13 to 20, characterized in that the supernatant is treated with a solvent / detergent. Process according to any one of claims 13 to 21, characterized in that the eluted immunoglobulins are concentrated by ultrafiltration and passed through conventional sterilization filtration, and then by filtration through nanometric filters with porosity decreasing from 100 to 15 nanometers. . Process according to any one of claims 13 to 22, characterized in that a pharmaceutically acceptable stabilizer is added to the concentrate and filtered immunoglobulin solution which is then packaged as a sterile solution and optionally frozen and lyophilized.