MXPA06000841A - Methods and compositions for increasing the efficiency of therapeutic antibodies using nk cell potentiating compounds - Google Patents

Abstract

The present invention relates, generally, to methods and compositions for increasing the efficiency of therapeutic antibodies. Their efficiency is enhanced through the increase of the ADCC mechanism. More particularly, the invention relates to the use of a therapeutic antibody in combination with compounds that block an inhibitory receptor or stimulate an activating receptor of an NK cell in order to enhance the efficiency of the treatment with therapeutic antibodies in human subjects.

Description

METHODS AND COMPOSITIONS TO INCREASE THE EFFICIENCY OF THERAPEUTIC ANTIBODIES THAT USE COMPOUNDS TO POTENTIATE NK CELLS FIELD OF THE INVENTION The present invention relates, in general, to methods and compositions for increasing the efficiency of therapeutic antibodies. More particularly, the invention relates to the use of a therapeutic antibody in combination with a compound that blocks an inhibitory receptor or stimulates a natural killer cell activating receptor, thereby enabling an enhancement of the cytotoxicity of the natural cytolytic cell in mammalian subjects in order to increase the efficiency of the treatment in human subjects, in particular through an increase of the ADCC mechanism.

BACKGROUND OF THE INVENTION Various therapeutic strategies in humans are based on the use of therapeutic antibodies. These include, for example, the use of therapeutic antibodies developed to decrease target cells, in particular diseased cells such as for example, virus-infected cells, tumor cells or other pathogenic cells. Typically these antibodies are monoclonal antibodies, of the IgG species, typically with human IgGl or IgG3 Fe portions. These antibodies may be natural or recombinant antibodies, and are often "humanized" mouse antibodies (eg, comprising functional domains from various species, typically an Fe portion of human or non-human primate origin, and with a variable region or a complementary determinant region (CDR) of mouse origin). Alternatively, the monoclonal antibody can be fully human through immunization in transgenic mice having the human Ig locus, or obtained through cDNA libraries derived from human cells. A particular example of these therapeutic antibodies is rituximab (Mabthera®, Rituxan®) which is a chimeric anti-CD20 monoclonal antibody constituted with human? And K constant regions (thus with the human Fe IgGl portion) bound to the domains with variable murine that confer a CD20 specificity. In recent years, rituximab had considerably modified the therapeutic strategy against malignancies. B lymphoproliferatives, in particular non-Hodgkin lymphomas (NHL). Other examples of humanized IgGl antibodies include alemtuzu ab (Campath-1H®) which is used in the treatment of B lymphocyte malignancies, and trastuzumab (Herceptin®) which is used in the treatment of breast cancer. Additional examples of therapeutic antibodies in development are discussed in the art. The mechanism of action of therapeutic antibodies is still a matter of debate. The injection of antibodies leads to the decrease of cells carrying the antigen specifically recognized by the antibody. This decrease can be delivered through at least three mechanisms: cellular cytotoxicity caused by antibodies (ADCC), complementary dependent lysis, and direct antitumor inhibition of tumor growth through the signals provided via the directed antigen by the antibody. While these antibodies represent a novel and efficient method for therapy in humans, in particular for the treatment of tumors, they do not always exhibit significant efficacy. For example, while rituximab, alone or in combination with chemotherapy, was shown to be effective in the treatment of both low and high-grade NHL, 30% to 50% of patients with low-grade NHL had no clinical response to rituximab. It has been suggested that the level of CD20 expression on lymphoma cells, the presence of a high tumor burden at the time of treatment, or low serum concentrations of rituximab may explain the lack of efficacy of rituximab in some patients. However, the real causes of treatment failure remain unknown. In addition, the use of therapeutic antibodies may be limited by side effects caused by their administration. For example, side effects such as, for example, fever, headaches, nausea, hypotension, wheezing, rashes, infections and many others can appear in patients, potentially limiting the possible amount of frequency with which antibodies can be administered. In this way, it would be very interesting to increase the efficiency of therapeutic antibodies, or be able to achieve therapeutic efficacy by using reduced doses of antibodies that are less likely to produce side effects. The present invention addresses these and other needs.

SUMMARY OF THE INVENTION The present invention sets forth novel methods for increasing the effectiveness of therapeutic antibodies. Without being limited by the following theory, it is believed that the surprising results achieved using the methods herein are the result of their ability to improve the ADCC mechanism in vivo, when therapeutic antibodies are injected. In fact, the present invention provides novel compositions and methods that overcome the current difficulty related to the efficiency of therapeutic antibodies. In the present invention it is shown that NK cells from an individual can have an ADCC supplied by therapeutic Ab (monoclonal antibody) deficient due to its lack of activation of NK cells, for example, by an inhibition of inhibitory receptors on cells NK Preferably, an increase in the ADCC mechanism is achieved by the administration of compounds that block an inhibitory receptor, or stimulate an activating receptor, on natural killer cells, thereby stimulating an enhancement of the cytotoxicity of natural killer cells in mammalian subjects. Preferably, the compound is an antibody or a fragment thereof. Antibodies or other compounds can be reacted with an inhibitor receptor of NK cells, for example, inhibitor receptor molecules for killing (KIR or NKG2A / C), or with activating receptors, for example, NCR such as, for example, NKp30, NKp44 or NKp46, on NK cells, thereby neutralizing the inhibition of the cells and increasing their ADCC activity. More specifically, the invention sets forth methods of treatment for a subject in which a compound, preferably an antibody or a fragment thereof, which blocks an inhibitory receptor or stimulates an activating receptor of an NK cell, is co-administered with the antibody. therapeutic to the subject. The inventors hereby demonstrate that the efficiency of a therapeutic antibody can be greatly improved by the co-administration, for example, co-injection, of this compound, preferably an antibody or a fragment thereof, which exceeds the inhibition of NK cells, for example, by blocking the inhibitory receptor or stimulating an activating receptor of an NK cell. The invention also relates to pharmaceutical compositions comprising a therapeutic antibody and a compound, preferably an antibody or a fragment thereof, that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell. The invention also relates to kits comprising a therapeutic antibody and a compound, preferably, an antibody or a fragment thereof, which blocks an inhibitory receptor or stimulates an activating receptor of an NK cell.

The invention also relates to the use of a compound, preferably an antibody or a fragment thereof, that blocks the inhibitory receptor or stimulates an activating receptor of an NK cell, to increase the efficiency of a treatment with a therapeutic antibody, or for increasing ADCC in a subject undergoing treatment with a therapeutic antibody. The invention also relates to the use of a compound, preferably an antibody or a fragment thereof, which blocks the inhibitory receptor or stimulates an activating receptor of an NK cell, and a therapeutic antibody for the preparation of a drug for the treatment of a disease. More particularly, the treatment of the disease requires the decrease of the target cells, preferably the diseased cells such as for example, virally infected cells, tumor cells or other pathogenic cells. Preferably, the disease is a cancer, an infectious or immune disease. More preferably, the disease is selected from the group consisting of a cancer, an auto-immune disease, an inflammatory disease and a viral disease. The disease is also related to a rejection of grafts, more particularly rejection of allografts, and a graft-versus-host disease (GVHD).

The present invention also encompasses a method for reducing the dosage of a therapeutic antibody, for example, an antibody that binds via an Fc receptor, preferably CD16 (FcγRIIIa, for its acronym in English). For example, the co-administration of a therapeutic antibody and a compound that blocks an inhibitory receptor or stimulates an activating receptor on NK cells that allows a lower dose of the therapeutic antibody to be used. These antibodies can be used at a dose of 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, than the recommended dose in the absence of the compound. In addition, the invention provides a method for determining a therapeutically effective reduced dosage of a therapeutic antibody, for example, an antibody bound by CDlβ, the method comprising i) co-incubating a first concentration of the therapeutic antibody with target cells and NK cells, and in the absence of a compound that blocks an inhibitory receptor or stimulates an activating receptor on NK cells; ii) co-subbing a second lower concentration of the therapeutic antibody with target cells, with NK cells, and in the presence of a compound that blocks an inhibitory receptor or stimulates an activating receptor on NK cells; iii) determining whether the decrease in target cells observed in step ii) is as great as the decrease observed in step i). If it is observed that step ii) is as effective as step i), then the relative concentrations of the compound and the therapeutic antibody may vary, and the observed decrease, in order to identify different conditions that may be suitable for use in a determined patient, for example, maximizing the decrease of target cells, the decreased dose of the therapeutic antibody, or the decreased dose of the compound, depending on the particular needs of the patient. In a particular aspect, the present invention provides a method of treating a disease in a human subject in need thereof, comprising: a) administering to the subject a compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell; and, b) administering to the subject a therapeutic antibody that can be bound by CD16. In one embodiment, the therapeutic antibody and the compound are administered in the subject simultaneously. In another embodiment, the compound is administered to the subject in approximately one week, approximately 4 days, approximately 3 days or the same day (e.g., in approximately 24 hours) of administration of the therapeutic antibody. In another modality, the disease is a cancer, an infectious or immune disease. In one embodiment, the method further comprises an additional step in which the activity or number of NK cells in the subject is assessed before or after administration of the compound. In another embodiment, the additional step involves i) obtaining NK cells from the subject prior to administration; ii) incubating the NK cells in the presence of one or more target cells that are recognized by the therapeutic antibody, in the presence or absence of the compound; and iii) assessing the effect of the compound on the ability of NK cells to decrease target cells; wherein a detection of that compound improves the ability of NK cells to decrease target cells indicating that the compound is suitable for use in the method, and that the method is suitable for use with the subject. In another aspect, the present invention provides a pharmaceutical composition comprising a therapeutic antibody, for example, that can be bound by CDlβ, to a compound that blocks an inhibitory receptor or stimulates an activating NK cell receptor, and a pharmaceutically acceptable carrier. In another aspect, the present invention provides a kit comprising a therapeutic antibody, for example, that can bind by CDlβ, to one or more compounds that block an inhibitory receptor or stimulate an activating receptor of NK cells. For any of the methods, compositions or equipment mentioned above, in one embodiment the therapeutic antibody has a human IgGl or an IgG3 Fe portion. In another embodiment, the compound is an antibody or a fragment thereof. In another embodiment, the therapeutic antibody is a monoclonal antibody or fragment thereof. In another embodiment, the therapeutic antibody is not conjugated with a radioactive or toxic entity. In another embodiment, the compound inhibits an inhibitor receptor of an NK cell. In another embodiment, the compound stimulates an activating receptor of an NK cell. In another embodiment, the compound is a human or humanized or chimeric antibody, or a fragment thereof. In one embodiment, the antibodies or therapeutic compounds can be antibody fragments or derivatives such as for example, among others, a Fab fragment, a Fab '2 fragment, a CDR and a ScFv. In one embodiment, the therapeutic antibody is a humanized or chimeric human antibody or a fragment thereof. In another embodiment, the therapeutic antibody is rituximab or Campath. In another embodiment, the antibody is rituximab, and the antibody is administered at a dosage lower than 375 mg / m2 per week. In another embodiment, the antibody is Campath, and the antibody is administered at a dosage lower than 90 mg per week. In one embodiment, the compound binds to at least one of the human NKG2, KIR2DL or KIR3DL receptors, and decreases the inhibition provided by NKG2, KIR2DL- or KIR3DL- related cytotoxicity of NK cells. In another embodiment, the compound blocks an inhibitory receptor of an NK cell selected from the group consisting of KIR2DL1, KIR2DL2 / 3, KIR2DL4, KIR2DL5A, KIR2DL5B, KIR3DL1, KIR3DL2, KIR3DL3, LILRB1, NKG2A, NKG2C NKG2E and LILRB5. In another embodiment, the compound binds to a common determinant of the human KIR2DL receptors and decreases the inhibition provided by KIR2DL- of the cytotoxicity of NK cells. In another embodiment, the compound binds to a common determinant of human receptors KIR2DL1, KIR2DL2, and KIR2DL3 and decreases the inhibition provided by KIR2DL1-, KIR2DL2-, and KIR2DL3- of the cytotoxicity of NK cells. In another embodiment, the compound inhibits the binding of an HLA-C allele molecule having a Lys residue at position 80 to a human KIR2DL1 receptor, and the binding of the HLA-C allele molecule has an Asn residue at the position 80 for the human receptors KIR2DL2 and KIR2DL3. In another embodiment, the compound binds to the same epitope as the monoclonal antibody DF200 produced by the hybridoma DF200. In another embodiment, the compound competes with the monoclonal antibody DF200 produced by the hybridoma DF200 by binding to a KIR receptor on the surface of a human NK cell. In another embodiment, the compound is a monoclonal antibody DF200 produced by the hybridoma DF200 or a fragment thereof. In one embodiment, the compound binds to a receptor selected from the group consisting of NKp30, NKp44, NKp46 and NKG2D. In another embodiment, the compound is derived or competes with a monoclonal antibody selected from the group consisting of AZ20, A76, Z25, Z231 and BAB281. In another aspect, the present invention provides a method for selecting a compound for administration in conjunction with a therapeutic antibody, the method comprising: i) providing a test compound that inhibits an inhibitory or stimulating receptor to an NK cell activating receptor;; ii) incubating the therapeutic antibody with the target cells specifically recognized by the therapeutic antibody in the presence of NK cells and in the presence or absence of the test compound; and iii) assessing the effect of the compound on the ability of NK cells to decrease target cells; wherein a detection that the compound improves the ability of NK cells to decrease target cells indicates that the compound is suitable for use in the method. In one embodiment, the compound improves the ability of the therapeutic antibody to kill target cells by 50%, 60%, 70%, 80%, 90%, 100%, 200%, 300%, 400%, 500%, or plus. In another embodiment, the compound is selected from the group consisting of an antibody, an antibody fragment, a monoclonal antibody, a fragment of a monoclonal antibody, a humanized antibody, a chimeric antibody, and a human antibody. In another embodiment, the target cells are cancer cells, virally infected cells, or cells affected by an auto-immune disorder. In another embodiment, the therapeutic antibody is rituximab or CAMPATH. In another aspect, the present invention provides a method for increasing the efficiency of a treatment involving the administration of a therapeutic antibody that can bind by CDlβ in a subject, the method comprising administering to the subject before, concurrent with, or after administration of the therapeutic antibody, a therapeutically effective amount of a compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell. In one embodiment, the compound increases the efficiency of the treatment by enhancing ADCC in the subject.

DESCRIPTION OF THE FIGURES Figure 1: The monoclonal antibody DF200 binds to a common determinant of various human KIR2DL receptors. Figure 2: reconstitution of lysis with a mAb (monoclonal antibody) anti-KIR2DL or on the C1R Cw4 bank at the effector / white ratio of 4/1. The monoclonal antibody DF200 decreases the inhibition provided by KIR2DL- of the positive NK cell cytotoxicity of KIR2DL1 (restorative lysis) on the positive Cw4 cells. Figure 3: Improvement of ADCC supplied by Rituxan of a positive NK clone KIR2DL1 on a positive EBV cell line Cw4 by blocking the KIR / HLA interaction. Cytolysis of NK clones carrying KIR2DL1 is tested against a transformed white cell line (CD20 positive) EBV positive of Cw4 at various effector / target ratios (from 1 to 4) in the presence of 5 μg / ml anti-CD20 antibody (Rutixan ) and 10 μg / ml of the EB6 antibody (anti-KIR2DL1); Rituxan alone; EBß only; or without any antibody. ADCC is greatly enhanced in the presence of the anti-KIR2DL1 antibody (EBß). Figure 4: Improvement of ADCC supplied by Campath or a positive NK clone KIR2DL1 in a positive EBV cell line Cw4 by blocking the KIR / HLA interaction. Cytolysis of the NK clone carrying KIR2DL1 is tested against a transformed white cell line (CD20 positive) EBV positive Cw4 in the presence of Campath and 100 μg / ml of the EBß antibody (anti-KIR2DL1); Campath alone; EBß only; or without any antibody. ADCC is greatly enhanced in the presence of the anti-KIR2DL1 antibody (EBß).

DETAILED DESCRIPTION OF THE INVENTION The present invention provides a method for increasing the efficiency of therapeutic antibodies. The invention more specifically states that the use of a compound, preferably an antibody or a fragment thereof, which potentiates NK cells, preferably by blocking an inhibitory receptor or activating an activating receptor of an NK cell, can significantly increase the efficiency of therapeutic antibodies. In fact, the inventors demonstrate that the efficiency of multiple therapeutic antibodies can be greatly improved by co-administration of an antibody directed against an NK cell receptor; for example, an inhibitory receptor. Therefore, the invention relates to a method for the treatment of a disease in a subject in need thereof comprising: a) administering to the subject a compound, preferably an antibody or a fragment thereof, that blocks a receptor inhibitor or stimulates an activating receptor of an NK cell; and, b) administering a therapeutic antibody to the subject. The therapeutic antibody can be bound by CDlβ of NK cells, preferably through its Fe region. Preferably, the therapeutic antibody has a human IgGl or a Fe IgG3 portion, in particular a monoclonal antibody or a fragment thereof, preferably in addition a humanized or chimeric humanized antibody or a fragment thereof, for example, rituximab. It is intended that compounds, preferably antibodies or a fragment thereof, that block the inhibitory receptor of an NK cell may be administered to the subject, before, simultaneously with or, after administration of the therapeutic antibody. The route of administration of the different antibodies depends on their bioavailability and pharmacokinetics. Preferably, the therapeutic antibody is administered within one week to the administration of the compounds, preferably the antibodies or a fragment thereof, which block the inhibitory receptor of an NK cell, more preferably in a period of about 5 or 2 days. Preferably, the therapeutic antibody is administered before or simultaneously with the compounds, preferably the antibodies or a fragment thereof, which block the inhibitory receptor of an NK cell. In an additional aspect, the invention relates to a method for increasing ADCC in a subject receiving a therapeutic antibody treatment, the method comprising administering to the subject before, simultaneously or after the administration of the therapeutic antibody an amount sufficient to increase ADCC of a compound, preferably an antibody or a fragment thereof, which blocks the inhibitory receptor of an NK cell. The therapeutic antibody can be bound by CDlβ on NK cells, preferably through its Fe region. Preferably, the therapeutic antibody has a human IgGl or an IgG3 Fe moiety, in particular a monoclonal antibody or a fragment thereof, of preferably further a human, humanized or chimeric antibody or a fragment thereof, for example, rituximab. In a further aspect, the invention relates to a method for increasing the efficiency of a treatment with therapeutic antibodies in a subject, the method comprising administering to the subject before, simultaneously or after the administration of the therapeutic antibody, an amount of a compound, preferably an antibody or a fragment thereof, which blocks the inhibitory receptor of an NK cell, sufficient to increase the efficiency of the therapeutic antibody. The therapeutic antibody can be bound by CDlβ, preferably through its Fe region. Preferably, the therapeutic antibody has a human IgGl or IgG3 Fe moiety, in particular a monoclonal antibody or a fragment thereof, in addition preferably a human antibody. humanized or chimeric or a fragment thereof, for example, rituximab.

DEFINITIONS As used herein, the following terms have the meanings attributed to them unless otherwise specified. In the sense in which it is used herein, "NK" cells refers to a sub-population of lymphocytes that is involved in an unconventional immunity. NK cells can be identified by virtue of certain characteristics and biological properties, such as, for example, the expression of specific surface antigens including CD16, CD56 and / or CD57, the absence of the TCR alpha / beta or gamma / complex. delta on the cell surface, the ability to bind to cytolytic cells that fail to express "by themselves" MHC / HLA antigens by activating specific cytolytic enzymes, the ability to kill tumor cells or other diseased cells that express a ligand for NK activating receptors, and the ability to release protein molecules called cytokines that stimulate or inhibit the immune response. Any of these features and activities can be used to identify NK cells, using methods well known in the art. The term "antibody", in the sense in which it is used herein, refers to polyclonal and monoclonal antibodies. Depending on the type of constant domain in the heavy chains, the antibodies are assigned to one of five major classes: IgA, IgD, IgE, IgG, and IgM. Several of these are further divided into sub-classes or isotypes, such as, for example, IgG1, IgG2, IgG3, IgG4, and the like. An exemplary immunoglobulin structural unit (antibody) comprises tetramer. Each tetramer is composed of two identical pairs of polypeptide chains, each pair having a "light" chain (approximately 25 kDa) and a "heavy" chain (approximately 50-70 kDa). The N-terminus of each chain defines a variable region of approximately 100 to 110 or more amino acids that are primarily responsible for the recognition of antigens. The terms variable light chain (VL, for its acronym in English) and variable heavy chain (VH, for its acronym in English) refer to these light and heavy chains respectively. The heavy chain constant domains corresponding to the different classes of immunoglobulins are referred to as "alpha", "delta", "epsilon", "gamma" and "mu" respectively. Subunit structures and three-dimensional configurations of different classes of immunoglobulins are well known. IgG and / or IgM are the preferred classes of antibodies used in this invention, with IgG being particularly preferred, because they are the most common antibodies in the physiological situation, because most of them are easily produced in a laboratory facility, and because IgG is specifically recognized by the Fe gammas receptors. Preferably, the antibody of this invention is a monoclonal antibody. Humanized, chimeric, human or otherwise suitable humanized antibodies are particularly preferred. Within the context of this invention, the term "therapeutic antibody or antibodies" more specifically designates any antibody that functions to decrease target cells in a patient. In particular, the therapeutic antibodies bind specifically to the antigens present on the surface of the target cells, for example, tumor-specific antigens present predominantly or exclusively in tumor cells. Preferably, the therapeutic antibodies include portions of human Fe, or are capable of acting on human Fe receptors. The therapeutic antibodies can be targeted to the target cells by any means, for example, ADCC or otherwise, and can be "stripped", ie, be without conjugated entities, or they can be conjugated with compounds such as for example, radioactive or toxins. The term "specifically binds to" means that an antibody can be preferentially bound in a competitive binding analysis for the binding partner, for example, an activating NK receptor such as, for example, NKp30, NKp4 or NKp46, or a human Fe gamma receptor, as assessed using any recombinant forms of the proteins, epitopes thereof, or natural proteins present on the surface of NK cells isolated or white relevant. Further described are further described and competitive binding analysis and other methods for determining specific binding are well known in the art. A "suitable human" antibody refers to any antibody, antibody derived, or antibody fragment that can be used in humans for example, in the therapeutic methods described herein. Suitable human antibodies include all types of humanized chimeric or fully human antibodies, or any antibodies in which at least a portion of the antibody is derived from humans or otherwise modified to avoid the immune response that is caused when they are used. natural non-human antibodies. By "immunogenic fragment", it is to be understood any polypeptide or peptide fragment that is capable of producing an immune response such as for example (i) the generation of antibodies that bind to the fragment and / or bind to any form of molecule that comprises the fragment, including the receptor bound to the membrane and the mutants derived therefrom, (ii) the stimulation of a T lymphocyte response involving the reaction of T lymphocytes with the bi-molecular complex comprising any MHC molecule and a derived peptide of the fragment, (iii) the binding of transfected vehicles such as, for example, bacteriophages or bacteria expressing genes encoding mammalian immunoglobulins. Alternatively, an immunogenic fragment also refers to any construct capable of producing an immune response as defined above, such as, for example, a peptide fragment conjugated to a carrier protein by covalent coupling, a chimeric recombinant polypeptide construct comprising the peptide fragment and its amino acid sequence, and specifically includes cells transfected with a cDNA of which the sequence comprises a portion encoding the fragment. For the purposes of the present invention, a "humanized" antibody refers to an antibody in which the constant and variable structural region of one or more human immunoglobulins is fused to the binding region, eg the CDR, of an animal immunoglobulin. . These humanized antibodies are designed to maintain the binding specificity of the non-human antibody from which the binding regions are derived, although to avoid an immunological reaction against the non-human antibody.

A "chimeric antibody" is an antibody molecule in which (a) the constant region, or a portion thereof, is altered, replaced or exchanged in such a way that the antigen binding site (variable region) is linked to a constant region of a different or altered class, an effector function and / or a species, or a totally different molecule that confers new properties to the chimeric antibody, for example, an enzyme, toxin, hormone, growth factor, drug, etc.; or (b) the variable region, or a portion thereof, is altered, replaced or exchanged with a variable region that has a different or altered antigenic specificity. In preferred embodiments of the present invention, nevertheless the chimeric antibody maintains the Fe region of the immunoglobulin, preferably a human Fe region, thereby allowing interactions with the human Fe receptors on the surface of target cells. Within the context of this invention, "enhanced", "active" or "activated" NK cells designate biologically active NK cells, more particularly NK cells that have the ability to lyse target cells. For example, an "active" NK cell is capable of killing cells that express an NK-activating receptor-ligand and fails to express "by itself" MHC / HLA antigens (cells incompatible with KIR). Examples of suitable target cells for use in re-directed killing assays are P815 and K562 cells, although any of several cell types can be used and are well known in the art (see, for example, Sivori et al. (1997). J. Exp. Med. 186: 1129-1136; Vitale et al. (1998) J. Exp. Med. 187: 2065-2072; Pessino et al. (1998) J. Exp. Med. 188: 953-960; Neri et al. (2001) Clin. Diag. Lab. Immun. 8: 1131-1135). "Enhanced" "active" or "activated" cells can also be identified by any other property or activity known in the art as associated with NK activity, such as, for example, the production of cytokines (eg, IFN-α). and TNF-a) of increases in free intracellular calcium levels. For the purposes of the present invention, "enhanced", "active" or "activated" cells particularly refer to NK cells in vivo that do not inhibit via stimulation to an inhibitory receptor, or in which this inhibition has been overcome, by example, via the stimulation of an activating receptor. As used herein, the term "activating NK receptor" refers to any molecule on the surface of NK cells that, when stimulated, causes a measurable increase in any property or activity known in the art according to the invention. it is associated with NK activity, such as, for example, the production of cytosines (for example, IFN-α and TNF-α), increases in intracellular free calcium levels, the ability to direct cells in a redirected extermination analysis according to it is described, for example, somewhere in the present specification, or the ability to stimulate NK cell proliferation. The term "activating KIR receptor" includes without limitation: NKp30, NKp44, NKp46, NKG2D, IL-12R, IL-15R, IL-18R and IL-21R. The term "activating NK receptor", in the sense in which it is used herein, excludes the IL-2 receptor (IL-2R). The methods for determining whether or not an NK cell is active or proliferating are described in greater detail below and are well known to those skilled in the art. In the sense in which it is used herein, the term "NK receptor", "to inhibit" or "inhibitor" refers to any molecule on the surface of NK cells that, when stimulated, causes a measurable decrease in any property or activity known in the art as associated with NK activity, such as, for example, the production of cytosines (for example IFN-α and TNF-α), increases in intracellular free calcium levels, or the ability to lyse target cells in a re-directed extermination analysis as described, for example, somewhere in the present specification. Examples of these receptors include KIR2DL1, KIR2DL2 / 3, KIR2DL4, KIR2DL5A, KIR2DL5B, KIR3DL1, KIR3DL2, KIR3DL3, LILRB1, NKG2A, NKG2C NKG2E and LILRB5. The methods for determining whether an NK cell is active or not are described in greater detail below and are well known to those skilled in the art. In the present invention, the term "blocking an inhibitory receptor or stimulating an activating receptor of an NK cell" refers to the ability of certain compounds, preferably antibodies, fragments or derivatives thereof, to interact preferably directly with at least an inhibitory or activating NK cell receptor, eg, KIR, NKG2A / C, NKp30, NKp44, NKp46 and others listed herein, and any neutralizing inhibitory signals of the receptor (in the case of inhibitory receptors) or stimulating signaling from of the receiver (in the case of activating receptors). With the inhibitory receptors, preferably the compound, preferably an antibody or a fragment thereof, is capable of blocking the interaction between HLA and the receptor. When the compound is an antibody, the antibodies can be polyclonal or, preferably, monoclonal. They can be produced by hybridomas or by recombinant cells subjected to technical studies to express the desired variable and constant domains. The antibodies can be single chain antibodies or other antibody derivatives that maintain the antigenic specificity and the lower region of articulation or a variant thereof such as for example a Fab fragment, a Fab '2 fragment, a CDR and a ScFv. These may be polyfunctional antibodies, recombinant antibodies, humanized antibodies or variants thereof. Within the context of this invention, a "common determinant" designates a determinant or epitope that is shared by various members of a group of related receptors, for example, the KIR2DL receptor group. The determinant or epitope may represent a peptide fragment or a conformational epitope shared by the members. In a specific embodiment, the common determinant comprises an epitope recognized by a monoclonal antibody DF200, NKVSF1 or EBß. Within the context of this invention, the term "antibody" that "binds" to a common determinant designates an antibody that binds to the determinant with specificity and / or affinity, for example, that does not bind essentially with high affinity or with specificity to others. unrelated motifs or a determinant or structures on the surface of human NK cells. More particularly, the binding of a monoclonal antibody according to this invention to the determinant can be discriminated from the binding of the antibody to another epitope or determinant. Compounds, preferably antibodies, capable of binding to NK cell inhibitory receptors and preventing their stimulation in this way are "neutralizing" or "inhibiting" compounds preferably antibodies, in the sense that they block, at least partially, the path of inhibitory signaling supplied by an inhibitor of NK cells, ie the KIR or NKG2A / C receptors. More importantly, this inhibitory activity can be exhibited with respect to various types of KIR or NKG2A / C receptors, such that these compounds, preferably antibodies, can be used in various subjects with high efficacy. The term "recombinant" when used with reference, for example, to a cell, or nucleic acid, protein or vector, indicates that the cell, nucleic acid, protein or vector, has been modified by the introduction of a heterologous nucleic acid or protein or the alteration of a natural nucleic acid or protein, or that the cell is derived from a cell modified in this way. Thus, for example, recombinant cells express genes that are not found within the natural (non-recombinant) form of the cell or natural genes are expressed that are otherwise expressed abnormally, sub-expressed or not expressed in full. Within the context of the present invention, a subject or patient includes any mammalian subject or patient, most preferably a human subject or patient.

THERAPEUTIC ANTIBODIES The present invention relates to the use of compounds to enhance NK cells together with therapeutic antibodies. Any of a wide variety of therapeutic antibodies can be used in the present invention. Essentially, any therapeutic antibody can be used, either "stripped" or conjugated with a radiolabel, toxin or other entity, either full length or a fragment; or either a true antibody or a modified derivative of an antibody. Preferably, the methods are used to enhance the efficacy of therapies in which the activity of NK cells plays a -not necessarily exclusive- role in the efficacy of the therapeutic antibodies administered, and preferably also the antibodies or fragments will naturally include, or will be modified to include a human Fe region or other domain that allows specific antibody recognition by human Fe receptors, e.g., Fe gamma receptors. The compounds of the present invention can be used to enhance the ability of therapeutic antibodies to decrease target cells that express an antigen that is specifically recognized by therapeutic antibodies. Accordingly, any disease or condition that is caused or aggravated at least in part by cells that can be targeted by a therapeutic antibody can be treated using the methods described herein. Specific examples of target cells include tumor cells, virus-infected cells, allogeneic cells, pathological immunocompetent cells (e.g., B lymphocytes, T cells, antigen presenting cells, etc.) involved in allergies, autoimmune diseases, allogeneic reactions , etc., or even healthy cells (for example, endothelial cells in an anti-angiogenic therapeutic strategy). The most preferred target cells within the context of this invention are tumor cells and cells infected by viruses. Therapeutic antibodies, for example, can provide a cytotoxic effect or cell lysis, in particular by antibody-dependent cell-supported cytotoxicity (ADCC). ADCC requires leukocyte receptors for the Fe of IgG (Fc? R) portion whose function is to unite the IgG sensitive a? Tigens for cytotoxic cells carrying Fc? R and to activate the activation machinery cell phone. Therefore, the therapeutic antibody is capable of forming an immunological complex. For example, an immune complex may be a targeted tumor covered by therapeutic antibodies. More particularly, the antibody can be bound by CDlβ, preferably through its Fe region. The determination that a therapeutic antibody binds or not to an Fc receptor? such as, for example, CDlβ can be titrated by any suitable form, for example by determining the binding with a recombinantly produced CDlβ polypeptide or a fragment thereof, optionally immobilized on a support, or for example by determining the binding of the therapeutic antibody to a cell of which is known or suspected to express CDlβ. The therapeutic antibodies may be polyclonal or, preferably, monoclonal. They can be produced by hybridomas or by recombinant cells subjected to technical studies to express the desired variable and constant domains. The antibodies can be single chain antibodies or other antibody derivatives that maintain the antigenic specificity and the smaller region of articulation or a variant thereof. They may be polyfunctional antibodies, recombinant antibodies, humanized antibodies, fragments or variants thereof. The fragment or a derivative thereof is preferably selected from a Fab fragment, a Fab '2 fragment, a CDR and a ScFv. Preferably, a fragment is an antigen binding fragment. Therapeutic antibodies comprising an antibody fragment may also include, but are not limited to: bi-specific antibodies; an example of a suitable bi-specific antibody comprises a binding region with antigen specific for CDlβ and an antigen binding region specific for a tumor antigen. Other antibody formats comprising fragments include derivatives of a recombinant bi-specific antibody that combine the binding regions of the two different antibodies on an individual polypeptide chain, also referred to as BiTEMR (Kufer P, et al TRENDS in Biotechnology 204; 22 ( 5): 238-244, and Baeuerle et al, Current Opinion in Molecular Therapeutics 2003; 5 (4): 413-419), the expositions thereof are incorporated herein by reference. Therapeutic antibodies are generally specific for surface antigens, for example, membrane antigens. The most preferred therapeutic antibodies are tumor-specific antigens (e.g., molecules specifically expressed by tumor cells), such as, for example, CD20, CD52, ErbB2 (or HER2 / Neu), CD33, CD22, CD25, MUC-1, CEA , KDR, aVß3, etc., in particular lymphoma antigens (e.g., CD20). Therapeutic antibodies preferably have an IgG1 or IgG3 Fe portion of human or non-human primate, more preferably human IgG1. In one embodiment, the antibodies will include modifications in their Fe moiety that enhance the interaction of the antibody with NK cells during ADCC. These modified therapeutic antibodies ("altered antibodies") generally comprise modifications, preferably, in the Fe region that modifies the binding affinity of the antibody to one or more FcγR. Methods for modifying antibodies with modified binding to one or more FcγR are known in the art, see, for example, PCT Publication Nos. WO 2004/016750 (International Application PCT / US2003 / 025399), WO 99/158572, WO 99/151642, WO 98/123289, WO 89/107142, WO 88/107089, and U.S. Patent Nos. 5,843,597 and 5,642,821 each are hereby incorporated by reference in their entirety. Therapeutic antibodies identified herein, such as, for example, D2E7 (Cambridge Antibody Technology Group, foot (Cambridge, UK) / BASF (Ludwigshafen, Germany)) used for the treatment of rheumatoid arthritis, or Infliximab (Centocor, Inc., Malvern, PA, used for the treatment of Crohn's disease and rheumatoid arthritis), or the antibodies set forth in International Patent Application PCT / US2003 / 025399 (which is incorporated herein by reference in its entirety) can be modified according to it was shown in the applications identified above and below and are used for the treatment of diseases for which these antibodies are typically used. In some embodiments, the invention provides altered antibodies having an altered affinity, an affinity either higher or lower, for an activating FcγR, for example, FcγRIII. In certain preferred embodiments, altered antibodies having higher affinity for FcγR are provided. Preferably, these modifications also have an effector function supplied by altered Fe. Modifications affecting effector function supplied by Fe are well known in the art (See, for example, U.S. Patent No. 6,194,351 which is incorporated herein by reference in its entirety). Amino acids that can be modified include, but are not limited to: proline 329, proline 331, and lysine 322. Proline 329 and / or 331 and lysine 322, can preferably be replaced with alanine, however, replacement with any other is also contemplated amino acid See International Publication No .: WO 00/142072 and U.S. Patent No. 6,194,551 which are incorporated herein by reference in their entirety. In this way, the modification of the Fe region may comprise one or more alterations for the amino acids found in the Fe region of the antibody. These alterations can result in an antibody with an effector function supplied by the altered antibody, an altered binding to other Fe receptors (e.g., Fe activation receptors), with altered ADCC activity, an altered Clq binding activity, an altered complement dependent on cytotoxicity activity, or any combination thereof. In one embodiment, the antibody is specifically recognized by a Fe gamma receptor such as for example FCGR3A (also referred to as CD16, FCGR3, immunoglobulin G receptor III.; IGFR3, receptor for the Fe fragment of IgG, low affinity Illa; see, for example, OMIM 146740), FCGR2A (also referred to as CD32, CDw32, receptor for the Fe fragment of IgG, low affinity lia, FCG2, immunoglobulin G receptor II, see, for example, OMIM 146790); FCGR2B (also referred to as CD32, receptor for the Fe fragment of IgG, low affinity Ilb, FCGR2B, FC-ga ma-RIIB, see, for example, OMIM 604590), FCG1RA (also called CD64, receptor for the Fe fragment of IgG, high affinity, IGFR1; see, for example, OMIM 146760); fragment FCGR1 of IgG, high affinity le, receptor IC Fe of immunoglobulin G IGFRC; see, for example, OMIM 601503); or FCGR1B (also referred to as CD64, receptor for the Fe fragment of IgG, high affinity Ib, receptor IB Fe of immunoglobulin G BIRF, IGFRB, see, for example, OMIM 601502). Typical examples of therapeutic antibodies of this invention are rituximab, alemtuzumab and trastuzumab. These antibodies can be used according to clinical protocols that have been authorized for use in human subjects. Additional specific examples of therapeutic antibodies include, for example, epratuzumab, basiliximab, daclizumab, cetuximab, labetuzumab, sevirumab, tuvurimab, palivizumab, infliximab, omalizumab, efalizumab, natalizumab, clenoliximab, etc. Optionally, when a compound that stimulates an activating receptor of an NK cell is a cytokine, the therapeutic antibody is an antibody other than rituximab or herceptin, or optionally other than an anti-CD20 or anti-HER2 / neu antibody. Other examples of preferred therapeutic antibodies for use in accordance with the invention include anti-ferritin antibodies (U.S. Patent Publication No. 2002/0106324), anti-pl40 and anti-sc5 antibodies (WO 02/50122), and antibodies anti-KIR (exterminating inhibitor receptor) (KIR receptors are described in Carrington and Norman, The KIR Gene Cluster, May 3, 2003, available at: http: // ww. ncbi. nlm. nih. gov / books), The expositions of each prior reference are incorporated herein by reference. Other examples of therapeutic antibodies are listed in the following table, any of which (and others) can be used in the methods herein. It will be appreciated, regardless of whether or not listed in the following table or described elsewhere in the present specification, any antibody that can decrease target cells, preferably by ADCC, can benefit from the methods herein, and the following Table 1 is not exclusive, neither with respect to the antibodies listed herein, nor with respect to the targets or indications of the antibodies that are listed. Table 1: Therapeutic antibodies COMPOUNDS TO REGULATE THE ACTIVITY OF NK CELLS The activity of NK cells is regulated by a complex mechanism that involves both stimulating and inhibitory signals. Accordingly, the therapy delivered by effective NK cells can be achieved by a stimulation of these cells or a neutralization of the inhibitory signals. It will be appreciated that any compound that has the effect of blocking, inhibiting or otherwise sub-regulating an NK cell inhibitory receptor, or of activating, stimulating, or otherwise promoting the activity or expression of an activating receptor of an NK cell. NK cells, it can be used. This includes compounds such as for example, cytokines, as well as, small molecules, polypeptides and antibodies that can bind to the receptors of NK cells and directly inhibit or stimulate it. It will also be appreciated that the mechanism by which the receptors are blocked or stimulated is not decisive for the advantages provided by the invention. For example, the compounds can increase the expression of an activating receptor, or inhibit the expression of an inhibitory receptor, the compounds can prevent interaction between a ligand and an inhibitory receptor or improve the interaction between a ligand and an activating receptor, or compounds can bind directly to the receptors and inhibit them (in the case of inhibitory receptors) or activate them (in the case of activating receptors). The decisive parameter is the effect that the compounds have on the ability of therapeutic antibodies to decrease their target cells in vivo. Any inhibitory receptor on the surface of an NK cell can be targeted by the compounds herein. NK cells are negatively regulated by a higher histocompatibility complex (MHC) class I-specific inhibitory receptors (Kárre et al., 1986; Ohlén et al, 1989; exposure thereof is incorporated in the present as reference). These specific receptors bind to polymorphic determinants of the highest histocompatibility complex (MHC), class I or HLA molecules and inhibit the lysis of natural cytolytic cells (NK), a family of receptors called groups for receptor recognition. similar to Ig exterminators (KIR) of class I alleles of HLA. There are several groups of KIR receptors, including KIR2DL, KIR2DS, KIR3DL and KIR3DS. KIR receptors having two Ig domains (KIR2D) identify the HLA-C allotypes: KIR2DL2 (formerly designated p58.1) or the closely related gene product KIR2DL3 recognizes an epitope shared by the allotype 2 HLA- (Cwl, 3, 7 and 8), whereas KIR2DL1 (p58.2) recognizes an epitope shared by the HLA-C allotypes of reciprocal group 1 (Cw2, 4, 5 and 6). Recognition by KIR2DL1 is regulated by the presence of a Lys residue at position 80 of the HLA-C alleles. The recognition of KIR2DL2 and KIR2DL3 is regulated by the presence of an Asn residue at position 80. Importantly, the vast majority of HLA-C alleles have either an Asn residue or a Lys residue at position 80. A KIR with three Ig domains, K1R3DL1 (p70), recognizes an epitope shared by the HLA-Bw4 alleles. Finally, a homodimer of molecules with three Ig domains KIR3DL2 (pl40), recognizes HLA-A3 and A-ll. Although KIR and other class I inhibitory receptors (Moretta et al, 1997, Valiante et al, 1997, Lanier, 1998, the exposure thereof is incorporated herein by reference) can be co-expressed by NK cells, in any repertoire NK of a given individual, there are cells that express an individual KIR and in this way, the corresponding NK cells are blocked only by cells expressing a group of specific class I alleles. Therefore, as will be described later, when inhibitory receptors are targeted, the methods of the present will often involve the administration of compounds that target multiple inhibitory receptors, thereby ensuring a broad-based effect reaching a maximum range of NK cells. In certain embodiments, the compound, preferably an antibody or a fragment thereof, blocks an inhibitor receptor of an NK cell, neutralizing the inhibitory signal of at least one inhibitory receptor selected from the group consisting of KIR2DL2, KIR2DL3, KIR2DL1, KIR3DL1, KIR3DL2, NKG2A and NKG2C. More preferably, the compound, preferably an antibody or a fragment thereof, which blocks the NK cell inhibitory receptor, is a compound, preferably an antibody or a fragment thereof, which neutralizes the inhibitory signal of KIR2DL2, KIR2DL3 and / or KIR2DL1. The invention also contemplates the use of a combination of various compounds, preferably antibodies or a fragment thereof, that blocks different NK cell inhibitory receptors. Preferably, the compounds, preferably antibodies or a fragment thereof, which block NK cell inhibitory receptors are specific for an inhibitory receptor selected from KIR2DL1, KIR2DL2, KIR2DL3, KIR3DL1, KIR3DL2, NKG2A and NKG2C and are capable of decreasing the inhibition provided by KIR- or NKG2A / C- related cytotoxicity of NK cells. For example, compounds that block NK cell inhibitory receptors may comprise an antibody having a specificity for KIR2DL1 and another having a specificity for KIR2DL2 and / or K1R2DL3. More preferably, the combination of compounds that block NK cell inhibitory receptors is able to decrease the inhibition provided by KIR2DL1-, KIR2DL2-, and KIR2DL3- of the cytotoxicity of NK cells. In other embodiments, a cocktail of one or more compounds that target one or more inhibitory receptors, as well as one or more compounds that target one or more activating receptors, will be administered. For example, monoclonal antibodies specific for KIR2DL1 have been shown to block alleles KIR2DL1 Cw4 (or the like) (Moretta et al., 1993; the presentation thereof is incorporated herein by reference). In another example, monoclonal antibodies against KIR2DL2 / 3 have also been described that block alleles KIR2DL2 / 3 HLAC w3 (or the like) (Moretta et al., 1993). Anti-NKG2A antibodies have been shown to block the inhibitory interaction between NKG2A and HLA-E. Optionally, the antibody can be selected from the group consisting of GL183 (KIR2DL2, L3, available from Immunotech, France and Beckton Dickinson, United States); EBß (KIR2DL1, available from Immunotech, France and Beckton Dickinson, United States.); AZ138 (KIR3DL1, available from Moretta et al, University of Genova, Italy); Q66 (KIR3DL2, available from Immunotech, France); Z270 (NKG2A, available from Immunotech, France); P25 (NKG2A / C, available from Moretta et al, University of Genova, Italy); and DX9, Z27 (KIR3DL1, available from Immunotech, France and Beckton Dickinson, United States). In a preferred aspect, the invention utilizes monoclonal antibodies, as well as, fragments and derivatives thereof, wherein the antibody, fragment or derivative cross-reacts with various KIR or NKG2A / C receptors on the surface of NK cells and neutralizes their inhibitory signals. In one embodiment, the invention utilizes a monoclonal antibody that binds to a common determinant of human KIR2DL receptors and inhibits the corresponding inhibitory pathway. Preferably, the invention utilizes a monoclonal antibody that binds to the KIR2DL1 and KIR2DL2 / 3 receptors on the surface of human NK cells and decreases the inhibition provided by KIR2DL1- and KIR2DL2 / 3- of the cytotoxicity of NK cells. The antibody specifically inhibits the binding of HLA-c molecules to the KIR2DL1 and KIR2DL2 / 3 receptors. Most preferably, the antibody facilitates the activity of NK cells in vivo. Because KIR2DL1 and KID2DL3 (or KIR2DL2) are sufficient to cover most of the HLA-C allotypes, the HLA-C allotypes of group 1 and the HLA-C allotypes of group 2 respectively, these antibodies can be used to increase the efficiency of a therapeutic antibody in the majority of human individuals, typically in approximately 90% of human individuals or more. In this embodiment, any of the antibodies described in PCT Patent Application No. PCT / FR 04/01702 filed on July 1, 2004, entitled "Compositions and methods for regulatory NK cell activity" may be used in accordance with the invention, the disclosure thereof is incorporated herein by reference. In a particular objective of this invention, the antibody that blocks the inhibitor receptor of an NK cell is a monoclonal antibody, wherein the antibody binds to a common determinant of human KIR2DL receptors and decreases the inhibition provided by KIR2DL- of the cytotoxicity of NK cells. The antibody binds more specifically to the same epitope as the monoclonal antibody DF200 or NKVSF1 produced by the hybridoma DF200 and NKVSF1 respectively and / or competes with the monoclonal antibody DF200 or NKVSF1 produced by the hybridoma DF200 and NKVSF1 respectively for binding to a KIR receptor. on the surface of a human NK cell. As discussed, examples of antibodies, functional analyzes and analyzes to determine whether or not antibodies compete for binding with antibodies are described in PCT Patent Application No. PCT / FR 04/01702. In a specific embodiment, the monoclonal antibody DF200 is the monoclonal antibody produced by the hybridoma DF200. In another embodiment, the monoclonal antibody is EB6, or the antibody binds to the same epitope as the monoclonal antibody EB6, or competes to bind with the monoclonal antibody EB6. In other embodiments, the antibody is a fragment or derivative of any of the antibodies DF200 or EB6. The hybridoma that produces the DF200 antibody has been deposited in the CNCM culture collection, as the identification number "DF200", registration number CNCM 1-3224, registered on June 10, 2004, Collection Nationale de Cultures de Microorganismos, Institut Pasteur, 25, Rué du Docteur Roux, F-75724 Paris Cedex 15, France. The NKVSF1 antibody is available from Serotec (Cergy Sainte-Christophe, France), catalog reference number MCA2243. In another embodiment of the present invention, the compound used to improve the efficacy of therapeutic antibodies stimulates an activating receptor of an NK cell. Any activating receptor can be used, for example, NKp30 (see, for example, PCT WO 01/36630, the disclosure thereof is hereby incorporated by reference in its entirety), NKp44 (see, for example, Vitale et al. (1998) J. Exp. Med. 187: 2065-2072, the disclosure thereof is hereby incorporated by reference in its entirety), NKp46 (see, for example, Sivori et al. (1997) J. Exp. Med. 186: 1129-1136; Pessino et al. (1998) J. Exp. Med. 188: 953-960; the expositions thereof are incorporated herein by reference in their totalities), NKG2D (see, for example, OMIM 602893), IL-12R, IL-15R, IL-18R, IL-21R, or an activating KIR receptor, for example, a KIR2DS4 receiver (Carrington and Norman, The KIR Gene Cluster, May 3, 2003, available at: http://www.ncbi.nlm.nih.gov/books), or any other receiver present in a fraction of NK cells, and whose activation leads to the activation or proliferation of the cell, preferably even if the cell had previously been inhibited via an inhibitory receptor such as, for example, an inhibitory KIR receptor. The compound can be any molecular entity, including polypeptides, small molecules, and antibodies. Exemplary compounds include any ligands, including natural, recombinant or synthetic ligands, that interact with the activating receptors. For example, a compound that stimulates an activating receptor of an NK cell may be a cytokine such as, for example, IL-12 which interacts with the IL-12 receptor (IL-12R), IL-15 which interacts with the IL receptor. -15 (IL-15R), IL-18 that interacts with the IL-18 receptor (IL-18R), IL-21 that interacts with the IL-21 receptor (IL-21R). These compounds are known, for example, from IL-12 (Research Diagnostics, NJ, DI-212), IL-15 (Research diagnostics, NJ, RDI-215), IL-21 (Asano et al., FEBS Lett.; 528: 70-6). Preferably, a compound that stimulates an activating receptor of an NK cell is a compound other than IL-2. Other example compounds that stimulate an activating receptor of an NK cell include antibodies that bind to an NK cell receptor selected from the group consisting of NKp30, NKp44, NKp46, NKG2D, KIR2DS4 and other KIR activators. In certain preferred embodiments, the activating receptor is a Natural Cytotoxicity Receptor (NCR) found in NK cells, preferably the NCR selected from the group consisting of group NKp30, NKp44 or NKp46, and the compound that stimulates an activating receptor, binds to the same epitope or competes to bind with any of the monoclonal antibodies selected from the group consisting of AZ20, A76, Z25, Z231 and BAB281. The binding of any compound to any of the NK cell receptors described herein can be detected using any of a variety of standard methods. For example, colorimetric ELISA tests, such as immunoprecipitation and radioimmunoassay, may be used. Competition analysis may be employed, for example, to compare the binding of a test compound to a known compound for binding to an NK cell receptor, in which the control (e.g., BAB281 that binds specifically to NKp46) and the test compounds are combined (or pre-adsorbed) and applied to a sample containing the protein containing the epitope, for example, NKp46 in the case of BAB281. Protocols based on ELISA, radioimmunoassay, Western blot, and the use of BIACORE are suitable for use in these simple competition studies and are well known in the art. The decrease in inhibition provided by KIR- or NKG2A / C- of the cytotoxicity of NK cells, or the stimulation of the activation provided by NKp30, NKp44, NKp46, or NKG2D- of the NK cells, can be assessed by various analyzes or tests, such as, for example, analysis of binding, cytotoxicity, and other molecular or cellular. In a specific variant, the inhibitory activity is illustrated by the ability of the compound, preferably an antibody, to reconstitute the KIR lysis or the positive NK NKG2A / C clones, respectively, on the positive HLA-C or HLA-E targets. . In another specific embodiment, the compound, preferably an antibody, is defined as the inhibitor of the binding of HLA-C molecules to the KIR2DL1 and KIR2DL3 receptors (or the closely related KIR2DL2), preferably further as their ability to alter the binding of a HLA-C molecule selected from Cwl, Cw3, Cw7, and Cw8 (or from an HLA-c molecule that tan an Asn residue at position 80) for KIR2DL2 / 3; and the binding of an HLA-C molecule selected from Cw2, Cw4, Cw5 and Cw6 (or from an HLA-c molecule having a Lys residue at position 80) for KIR2DL1. The inhibitory or enhancing activity of a compound of this invention, preferably an antibody, can be assessed in any of several ways, for example, by its effect on intracellular free calcium as described, for example, in Sivori et al. (1997) J. Exp. Med. 186: 1129-1136, the disclosure thereof is incorporated herein by reference. The activity of NK cells can also be assessed using cell-based cytotoxicity assays, for example, by measuring the release of chromium, such as, for example, by assessing the ability of the antibody to stimulate NK cells to kill target cells such as example, P815 cells, K562, or suitable tumor cells as set forth in Sivori et al. (1997) J. Exp. Med. 186: 1129-1136; Vítale et al. (1998) J. Exp. Med. 187: 2065-2072; Pessino et al. (1998) J. Exp.

Med. 188: 953-960; Neri et al. (2001) Clin. Diag. Lab. Immun. 8: 1131-1135); Pende et al. (1999) J. Exp. Med. 190: 1505-1516), the total exposures of each of them are incorporated herein by reference. Suitable cytotoxicity analyzes are also provided in the examples section of the present specification. In a preferred embodiment, the antibodies cause at least a 10% increase in NK cytotoxicity, preferably at least a 40% or 50% increase in NK cytotoxicity, or more preferably at least a 70% increase in NK cytotoxicity. NK cytotoxicity. The activity of NK cells can also be directed to the use of an analysis for cytokine release, wherein the NK cells are incubated with the antibody to stimulate the production of cytokines of NK cells (for example, the production of IFN-α and TNF-a). In an example protocol, the production of IFN-? from PBMC is assessed by the cell surface in intracytoplasmic staining and analysis by flow cytometry after 4 days of culture. Briefly, Brefeldin A (Sigma Aldrich) is added at a final concentration of 5 μg / ml during the last 4 hours of culture.

The cells are then incubated with anti-CD3 mAb and anti-CD56 before permeabilization (IntraPrep ™; Beckman; -Coulter) and stained with PE-anti-IFN-? or PE-IgGI (Pharmingen). The production of GM-CSF and IFN-α was measured from polyclonal activated NK cells in supernatants using ELISA (GM-CSF: DuoSet Elisa, R &D Systems, Minneapolis, MN; IFN- ?: OptElA set, Pharmingen). In a preferred embodiment, the ability of the antibody to activate human NK cells is evaluated, wherein the ability to activate human NK cells as well as non-human NK cells indicates that the compounds are suitable for use in the present invention. In particular, the ability of the compound to improve the ability of therapeutic antibodies to direct the decrease of suitable target cells by NK cells in vitro or in vivo can be assessed. The compounds of this invention, preferably antibodies, may exhibit partial inhibitory or stimulating activity, for example, partially reducing the inhibition provided by KIR2DL- of the cytotoxicity of NK cells, or partially activating an NK cell through any level of stimulation of NCR or other receivers. More preferred compounds are capable of inhibiting (or stimulating, in the case of activating receptors) at least 20%, preferably at least 30%, 40% or 50% or more of the activity of NK cells, for example, as measured in an analysis for cellular toxicity, compared to cells in the absence of the compound.

Also, the preferred compound can provide an increase in the decrease of target cells by 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 200%, 300%, 400%, 500%, 1000%, or more in relation to the level of decrease in absence of the compound. Alternatively, the preferred compounds of this invention, preferably antibodies, are also capable of inducing lysis of a matched, compatible or autologous white cell population with HLA, i.e., the cell population that could not be effectively used by NK cells in the absence of the antibody. Accordingly, the compounds of this invention can also be defined as facilitators of NK cell activity in vivo. The invention also contemplates embodiments in which the compounds that stimulate activating receptors, or, preferably, block the inhibitory receptor of an NK cell, are fragments of this monoclonal antibody that have practically the same antigenic specificity, including, without limitation: a fragment Fab, a Fab '2 fragment, a CDR, a ScFv. In addition, the monoclonal antibody can be humanized, human, or chimeric (e.g., a bi-specific antibody or functional groups). While the antibodies that stimulate the activating receptors can also be fragments, they preferably have total length. Derivatives, for example, modified or conjugated heterologous functional groups or other compounds, can be used for any of the antibodies described herein. Antibodies that block the inhibitory receptor or stimulate an activating receptor of an NK cell according to the invention can be produced by a variety of techniques known in the art. Typically, they are produced by immunizing a non-human animal with an immunogen comprising a KIR polypeptide, NKG2A / C, NCR (eg, NKp30, NKp44, NKp46), or an immunogenic fragment of any of the NKG2D polypeptides, and a collection of spleen cells (to produce hybridomas by fusion with suitable cell lines). Methods for producing monoclonal antibodies from various species are well known in the art (see, for example, Harlow et al., "Antibodies: A Laboratory Manual," CSH Press, 1988; Goding, "Monoclonal Antibodies: Principles and Practice, "Ademic Press, 1986, the expositions of which are incorporated herein by reference). More specifically, these methods comprise immunizing a non-human animal with the antigen, followed by recovery of spleen cells which are then fused with immortalized cells, such as, for example, myeloma cells. The resulting hybridomas produce the monoclonal antibodies and can be selected by limiting dilutions to isolate individual clones. The antibodies can also be produced by the selection of combinatorial libraries of immunoglobulins, as described for example in Ward et al (1989); the disclosure thereof is incorporated herein by reference. Preferred antibodies that block the inhibitory receptor or stimulate an activating receptor of an NK cell according to the invention are prepared by immunization with an immunogen comprising an activating or inhibitory NK cell receptor, eg, a KIR2DL polypeptide, more preferably a human KIR2DL polypeptide. The KIR2DL polypeptide may comprise the full length sequence of a human KIR2DL polypeptide, or a fragment or derivative thereof, typically an immunogenic fragment, ie, a portion of the polypeptide comprising an epitope, preferably an epitope of T or B lymphocytes. These fragments typically contain at least 7 consecutive amino acids of the mature polypeptide sequence, even more preferably at least 10 consecutive amino acids thereof. They are derived essentially from the extracellular domain of the receptor. In a preferred embodiment, the immunogen comprises wild type human KIR2DL, NCR, or other polypeptide in a lipid membrane, typically on the surface of a cell. In a specific embodiment, the immunogen comprises intact NK cells, in particular human NK cells intact, treated or optionally used. While therapeutic antibodies may have modified Fe regions to enhance their binding by receptors such as, for example, CDlβ, in certain embodiments the NK cell enhancing antibodies will have altered Fe regions to reduce their affinity for Fe receptors, thereby reducing the probability that the NK cells bound by the antibodies by themselves bind and lyse. Antibodies that block receptors KIR2DL of NK cells can be produced by methods comprising: i) immunizing a non-human mammal with an immunogen comprising a KIR2DL polypeptide; ii) preparing monoclonal antibodies from the immunized animal, wherein the monoclonal antibodies bind to the KIR2DL polypeptide; iii) selecting the monoclonal antibodies from step ii) that cross-react with at least two different serotypes of K1R2DL polypeptides; and iv) selecting the monoclonal antibodies of (c) that decrease the inhibition provided by KIR2DL of the NK cells. The order of steps (iii) and (iv) can be changed. Optionally, the method may further comprise additional steps for the preparation of fragments or derivatives of the monoclonal antibody, as will be described below. In another variant, the method comprises: i) selecting, from a library or repertoire, a monoclonal antibody or a fragment or derivative thereof cross-reactive with at least two different serotypes of KIR2DL polypeptides; and selecting an antibody from step i) that decreases the inhibition provided by KIR2DL of the NK cells. It will be appreciated that any of these methods may be used for the selection of any antibodies or antibody fragments that are specific for any group of NK cell receptors (inhibitors or activators) that share one or more epitopes. For example, similar methods can be used for the preparation of antibodies that block a KIR3DL or an NKG2A / C NK cell receptor, or stimulate an NK cell activating receptor. In a preferred embodiment, the non-human animals used in these methods, or used in the production of any of the antibodies described herein, is a mammal, such as, for example, a rodent (eg, mouse, rat, etc.). ), bovine, porcine, equine, rabbit, goat, sheep, etc. Also, any of the antibodies described herein can be genetically modified or subjected to technical studies that are suitable for human beings, for example, humanized, chimeric, or human antibodies. Methods for humanizing antibodies are well known in the art. In general, a humanized antibody according to the present invention has one or more amino acid residues introduced therein from the original antibody. These murine or other non-human amino acid residues are often referred to as "import" residues, which are typically extracted from a "import" variable domain. Humanization can be performed essentially following the method of Winter et al. (Jones et al., (1986) Nature 321: 522; Riechmann et al. (1988) Nature 332: 323; Verhoeyen et al. (1988) Science 239: 1534 (1988)). In some cases, these "humanized" antibodies are chimeric antibodies (Cabilly et al., U.S. Patent No. 4,816,567), wherein substantially less than an intact human variable domain has been replaced by the corresponding sequence of the original antibody. In practice, humanized antibodies according to this invention are typically human antibodies in which some CDR residues and possibly some FR residues are replaced by residues from analogous sites in the original antibody. Another method for producing "humanized" monoclonal antibodies is to use a XenoMouse® (Abgenix, Fremont, CA) as the mouse used for immunization. A XenoMouse is a murine host that has had its immunoglobulin genes replaced by functional human immunoglobulin genes. In this way, the antibodies produced by this mouse or in hybridomas produced from B lymphocytes of that mouse, are already humanized. The XenoMouse is described in U.S. Patent No. 6,162,963 which is incorporated herein by reference in its entirety. An analogous method can be achieved using a HuMAb-Mouse ™ (Medarex). Human antibodies can also be produced according to various other techniques, such as, for example, when using, for immunization, other transgenic animals that have undergone technical studies to express a repertoire of human antibodies (Jakobovitz et al., Nature 362 (1993) 255), or by selecting repertoires of antibodies using methods that display phages. These techniques are known to the experts and can be implemented by starting from monoclonal antibodies as set forth in the present application. The antibodies of the present invention can also be derived to "chimeric" antibodies (immunoglobulins) in which a portion of the heavy and / or light chain is identical or homologous with the corresponding sequences in the original antibody, while the rest of the chains are identical or homologous with the corresponding sequences in the antibodies derived from other species or belonging to another class or subclass of antibodies, as well as the fragments of these antibodies, as long as they exhibit the desired biological activity (Cabilly et al., supra; Morrison et al., (1984) Proc. Nati. Acad. Sci. 81: 6851). It will also be appreciated that when the compound is blocked to the inhibitory receptor of an NK cell or stimulates an activating receptor of an NK cell is an antibody, this antibody can be polyclonal or, preferably, monoclonal. The antibody can be produced by a hybridoma or by a recombinant cell subjected to technical studies to express the desired variable and the constant domains. The antibody can be an individual chain antibody or other antibody derivative that maintains the antigenic specificity and the lower region of articulation or a variant thereof. The antibody can be a polifunctional antibody, a recombinant antibody, a humanized antibody, or a fragment or derivative thereof. The fragment derived therefrom is preferably selected from a Fab fragment, a Fab '2 fragment, a CDR and a ScFv. Preferably, a fragment is a fragment binding with antigen. An antibody comprising an antibody fragment can also include, but is not limited to, bi-specific antibodies. An example is a bi-specific antibody comprising an antigen-binding region specific for an activating receptor and an antigen-binding region specific for a tumor antigen (see, PCT Publication No. WO 01/71005), they are incorporated herein by reference).

COMPOSITION AND ADMINISTRATION The invention relates to a composition comprising at least one compound, preferably an antibody or a fragment thereof, which blocks the inhibitory receptor or stimulates an activating receptor of an NK cell, and a therapeutic antibody, the use of the composition for increasing the efficiency of the therapeutic antibody, for increasing ADCC in a subject treated with a therapeutic antibody, or for the treatment of a subject having a disease, more particularly a disease requiring the decrease of target cells, preferably diseased cells such as, for example, virally infected cells, tumor cells or other pathogenic cells. Preferably, the disease is selected from the group consisting of a cancer, an auto-immune disease, an inflammatory disease, a viral disease. The disease is also associated with a rejection of grafts, more particularly a rejection of allografts, and a graft-versus-host disease (GVHD). More particularly, the treatment of the disease requires the decrease of target cells, preferably the diseased cells such as for example, virally infected cells, tumor cells or other pathogenic cells. Preferably, the disease is a cancer, an infectious or immune disease. More preferably, the disease is selected from the group consisting of a cancer, an auto-immune disease, an inflammatory disease, a viral disease. The disease is also associated with a rejection of grafts, more particularly rejection of allografts, and graft-versus-host disease (GVHD). These diseases include the neoplastic proliferation of hematopoietic cells. Optionally, the diseases are selected from the group consisting of lymphoblastic leukemia, acute or chronic myelogenous leukemia, Hodgkin's lymphoma, non-Hodgkin's lymphoma, myelodysplastic syndrome, multiple myeloma, and chronic lymphocytic leukemia. These diseases also include ENT cancer, colorectal cancers, breast cancer, epithelial cancer. These diseases include CMV infection, and hepatitis B. The diseases include Crohn's disease, rheumatoid arthritis, asthma, psoriasis, multiple sclerosis or diabetes. In particular, any disease listed in the table provided below can be treated. The therapeutic antibody can be bound by CD16, preferably through its Fe region. Preferably, the therapeutic antibody has an Fe-portion of IgG1 or a human IgG3, in particular a monoclonal antibody or a fragment thereof, further preferably a human, humanized or chimeric antibody or a fragment thereof, for example, rituximab. The compound, preferably an antibody or fragment thereof, which blocks the inhibitory receptor or stimulates the activating receptor of an NK cell binds to at least one of the human KIR, NKG2A / C, NCR or NKG2D receptors, and decreases the inhibition provided by KIR2DL, KIR3DL and / or NKG2A / C of the cytotoxicity of NK cells, or stimulates the activation provided by NCR or NKG2D of the cytotoxicity of NK cells. In a preferred embodiment, a human KIR2DL receptor is used, for example a receptor selected from the group consisting of human receptors KIR2DL1, KIR2DL2, KIR2DL3 or a human KIR3DL receptor is used, for example, a receptor selected from the group consisting of KIR3DL1 and KIR3DL2. In a preferred embodiment, the NK cell enhancing compound binds to at least one of the human KIR2DL receptors and decreases the inhibition provided by related KIR2DL of the cytotoxicity of NK cells. Preferably, the human receptor KIR2DL is selected from the group consisting of the human receptors KIR2DL1, KIR2DL2, KIR2DL3. In a preferred embodiment, the compound, preferably an antibody or a fragment thereof, binds to a common determinant of the human KIR2DL receptors and decreases the inhibition provided by KIR2DL of the cytotoxicity of NK cells. More preferably, the compound, preferably an antibody, binds to a common determinant of human receptors KIR2DL1, KIR2DL2, KIR2DL3 and decreases the inhibition provided by KIR2DL1, KIR2DL2, KIR2DL3 of NK cell cytotoxicity. In a particular embodiment, the compound, preferably an antibody, inhibits the binding of a molecule of the HLA-C allele having a Lys residue at position 80 to a human KIR2DL1 receptor, and the binding of a molecule of the HLA-C allele. which has an Asn residue at position 80 with the human KIR2DL2 and KIR2DL3 receptors. In another particular embodiment, this antibody binds to the same epitope as the monoclonal antibody DF200 produced by the hybridoma DF200. Optionally, this antibody competes with the monoclonal antibody DF200 produced by the hybridoma DF200 for binding to a KIR receptor on the surface of a human NK cell. In a preferred embodiment, the antibody is the monoclonal antibody DF200 produced by the hybridoma DF200. In another embodiment, the antibody competes or binds to the same epitope as the monoclonal antibody EB6. The composition according to the present invention may comprise a combination of various compounds, preferably antibodies or a fragment thereof, that block different NK cell inhibitory receptors, and / or stimulate one or more activating receptors of NK cells. Preferably, the compounds, preferably antibodies or a fragment thereof, which blocks the NK cell inhibitory receptors are specific for an inhibitory receptor selected from KIR2DL1, KIR2DL2, KIR2DL3, KIR3DL1, KIR3DL2, NKG2A and NKG2C, and are capable of of decreasing the inhibition provided by KIR or NKG2A / C related cytotoxicity of NK cells. More preferably, the combination of the "neutralizing" compounds is capable of decreasing the inhibition provided by KIR2DL1, KIR2DL2, and KIR2DL3 of the cytotoxicity of NK cells. By providing a combination of compounds, a maximum number of different inhibitory receptors will be blocked in a maximum number of patients. Also, combinations of compounds that stimulate different activating compounds (or, as with inhibitory receptors, bind to different epitopes within a single receptor), can be used, for example, compounds that together lead to the activation of any combination of two or more receptors selected from the group consisting of NKp30, NKp44, NKp46, and NKG2D. Also, combinations comprising one or more compounds that block an inhibitory receptor, and one or more compounds that stimulate an activating receptor, can be used. As will be described below, in a preferred embodiment, a sample of NK cells from a patient can be obtained prior to the application of the methods herein, and the receptivity of the NK cells to different combinations of compounds can be assessed, for example, in the presence of target cells and the therapeutic antibody.

The compositions of this invention may comprise any pharmaceutically acceptable carrier or excipient, typically a buffer, isotonic solutions, aqueous suspensions, optionally supplemented with stabilizing agents, preservatives, etc. Typical formulations include a saline solution and, optionally, a protective or stabilizing molecule, such as, for example, a high molecular weight protein (e.g., human serum albumin). Also provided are kits comprising any combination of one or more therapeutic antibodies, one or NK cell enhancing compounds, and, preferably, instructions for their use. According to the methods and compositions of the present invention, the compounds, preferably an antibody or a fragment thereof, that block an inhibitory receptor or stimulate an activating receptor of an NK cell and therapeutic antibodies are administered in an "efficient" amount. or "therapeutically effective". The efficient amount of therapeutic antibodies administered to the container may be between about 0.1 mg / kg to 20 mg / kg. The efficient amount of the antibody however depends on the form of the antibody (whole Ig, or fragment), the affinity of the mAb and the pharmacokinetic parameter that must be determined for each of the particular antibodies. The efficient amount of compounds, preferably an antibody or a fragment thereof, which blocks the inhibitory receptor or stimulates an activating receptor of an NK cell administered to the container may be between about 0.1 mg / kg to 20 mg / kg. The efficient amount of the antibody however depends on the form of the antibody (whole Ig, or fragments), the affinity of the mAb and the pharmacokinetic parameters that must be determined for each of the particular antibodies. In an important embodiment of the invention, the use of the compounds herein may allow a therapeutic efficacy to be achieved with reduced doses of therapeutic antibodies. The use (for example, the dosage regimen, administration) of therapeutic antibodies can be limited by side effects, for example, in the case of rituximab, fever, headaches, wheezing, drop in blood pressure, and others. Accordingly, while in many patients a standard dosage of the therapeutic antibodies will be administered together with the NK cell enhancing compounds described above (ie, the recommended dose in the absence of any other compounds), thereby improving the efficiency of the dosage standard in patients who need even greater therapeutic efficiency, in other patients, for example, those severely affected by side effects, the administration of the compounds of the present will allow a therapeutic efficiency to be reached at a reduced dosage of therapeutic antibodies, avoiding with this side effects. In practice, a skilled physician will be able to determine the ideal dosage and the administrative regimen of the therapeutic antibody and the NK cell enhancing compound for a given patient, for example, the therapeutic strategy that should be suitable in view of the particular needs and the overall condition of the patient. Many references are available for guidance in determining suitable dosages, both for therapeutic antibodies and for NK cell enhancing compounds, for example, Remington: The Science and Practice of Pharmacy, by Gennaro (2003), ISBN: 0781750253; Goodman and Gilmans The Pharmacological Basis of Therapeutics, by Hardman, Limbird & Gilman (2001), ISBN: 0071354697; Rawlins E. A., editor, "Bentley's Textbook of Pharmaceutics", London: Bailliere, Tindall and Cox, (1977); and others. In one embodiment, a physician can gradually decrease the amount of the therapeutic antibody provided in conjunction with the administration of any NK cell enhancing compounds present.; either in the terms of dosage or frequency of administration, and to monitor the efficacy of the therapeutic antibody; for example, monitor the activity of NK cells; monitor the presence of target cells in the patient, monitor various clinical indications, or by any other means, and, in view of the results of monitoring, adjust the relative concentrations or modes of administration of the therapeutic antibodies and / or the compound NK cell enhancer to optimize the therapeutic efficiency and the limitation of side effects. In another set of modalities, NK cells will be obtained from the patient prior to the administration of the therapeutic antibody and the NK cell enhancing compounds (and, if appropriate, during treatment), and will be titrated to determine the ideal compound or adequate of the compounds that will be used for maximum effectiveness. For example, the identity of the inhibitory or activating receptors present in NK cells can be determined, and the administered compounds that specifically target the most prominent receptors.

Alternatively, the NK cells obtained can be incubated with the therapeutic antibody and the target cells, and the ability of one or more compounds to intensify the decrease of target cells can be assessed. Then any of one or more compounds that are more effective in improving in vitro decline can be selected for use in the present methods of treatment. Suitable dosages of the compounds and / or therapeutic antibodies which in general can also be determined in vitro or in animal models, for example, in vitro by incubating various concentrations of a therapeutic antibody in the presence of target cells, NK cells (preferably cells Human NK), optionally other immune cells, and variable concentrations of one or more NK cell enhancing compounds, and assessing the degree or rate of decrease of target cells under various conditions, using standard analysis (e.g., as described in the section of examples). Alternatively, variable dosages of the therapeutic antibodies can be administered to animal models for diseases that can be treated with the antibodies (e.g., an animal model for NHL in the case of rituximab), together with variable dosages of the compounds described herein. , and the effectiveness of the antibodies (for example, as determined by any suitable clinical, cellular, or molecular analysis or criteria) in the treatment of animals that may be assessed. The composition according to the present invention can be injected directly to a subject, typically by the intravenous, intraperitoneal, intra-arterial, intramuscular or transdermal route. It has been shown that various monoclonal antibodies will be efficient in clinical situations, such as for example, Rituxan (Rituximab) or Xolair (Omalizumab), and in similar administration regimens (ie, formulations and / or dosages and / or administration protocols) they can be used with the composition of this invention. In addition, the compositions of this invention may further comprise or may be used in combination with other active agents or therapeutic programs such as, for example, chemotherapy or other immunotherapies, either simultaneously or sequentially. In a certain preferred example, the method of the invention further comprises one or more injections of two or more compounds that block an inhibitory receptor or stimulate an activating receptor of an NK cell. In this way, these two or more compounds can be used in combination. This can serve to cause an even greater increase in ADCC and the efficiency of therapeutic antibodies, and / or can serve to reduce the dosage of a particular compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell. For example, it is known that compounds such as, for example, IL-2 are toxic at increased dosages. Therefore, the invention preferably provides a method for treating a disease in a subject in need thereof comprising: a) administering to the subject at least two compounds, preferably an antibody or a fragment thereof, that blocks a inhibitor receptor or stimulate an activating receptor of an NK cell; and b) administering a therapeutic antibody to the subject. For example, a preferred regimen is when the two compounds are (i) a first compound selected from the group consisting of an antibody that stimulates an NCR or NKG2D receptor or an activating KIR receptor, and an antibody that blocks an inhibitory KIR receptor or NKG2A , and (ii) a second compound selected from the group consisting of IL-12, IL-15, IL-18 and IL-21. Therefore the invention also provides a method for the treatment of a disease in a subject in need thereof comprising: a) administering to the subject a compound according to the invention, preferably an antibody or a fragment thereof, which blocks an inhibitory receptor or stimulate an activating receptor of an NK cell; and b) administering a therapeutic antibody to the subject; and c) administering the subject IL-2. IL2 which is available from Research Diagnostics, NJ, RDI-202, or Chiron Corp. (Emeryville, CA). The cytokines can be administered in accordance with any suitable administration regimen, and can be administered before, simultaneously and / or after the administration of the compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell, and before, simultaneously and / or after administration of the therapeutic antibody. In a typical example, the cytokine is administered daily for a period of 5-10 days, the cytokines will be injected first on the same day as the first injection of the compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell. Preferably the method comprises one or two injections / day of cytokines subcutaneously. The dosage of the cytokine will be selected depending on the condition of the patient to be treated. In preferred examples, a relatively low dose of the cytokine can be used. For example, an effective dose of a is typically less than 1 million units / square meters / day of cytokines, when the pharmaceutical composition containing cytokines is used for a daily subcutaneous injection. In a preferred example, IL-2 is injected subcutaneously at daily doses below 1 million units / m2 for 5 to 10 days. The use of cytokines in greater detail is described in International Patent Publication No. PCT / EP / 0314716 and U.S. Patent Application No. 60 / 435,344 entitled "Pharmaceutical Compositions Having an Effect on the Proliferation of NK Cells and a method using the same "the expositions thereof are incorporated herein by reference. It will also be appreciated that therapeutic antibodies and NK cell enhancing compounds can be co-administered, e.g., co-injected, or can be administered simultaneously although in different formulations, or can be administered independently, e.g., the compound is administered hours, days, or weeks before or after administration of the compound. The additional aspects and advantages of this invention will be set forth in the following experimental section, which should be interpreted as illustrative and not limiting of the scope of this application.

EXAMPLES Example 1: Generation of a concentrated KIR2DL antibody by evaporation Purification of PBL and generation of polyclonal or clonal NK cells PBL are derived from healthy donors by Ficoll Hypaque gradients and the decrease of plastic adherent cells. To obtain enriched NK cells, the incubated PBL were incubated with the anti-CD3, anti-CD4 and anti-HLA-DR mAbs (30 mns C at 4 ° C), followed by goat anti-mouse magnetic beads (Dynal) ( 30 mns at 4 ° C) and immunomagnetic selection by methods known in the art (Pende et al., 1999). CD3 negative, CD4 negative, negative DR cells are cultured in irradiated feeder cells and 100 U / ml interleukin 2 (Proleukin, Chiron Corporation) and 1.5 ng / ml of phytohemagglutinin A (Gibco BRL) to obtain polyclonal NK cell populations. The NK cells are cloned by limiting the dilution and the clones of NK cells are characterized by flow cytometry for the expression of cell surface receptors. In this study the following clones were used: CPU, CN5 and CN505 are positive KIR2DL1 clones and stained by EB6 or XA-141 antibodies. CN12 and CP502 are positive KIR2DL3 clones and stained by the GL183 antibody.

Analysis by flow cytometry The mAbs used were produced in the laboratory JT3A (IgG2a, anti-CD3), EB6 and GL183 (IgGl anti-KIR2DL1 and KIR2DL3 respectively), XA-141 IgM anti-KIR2DL1 (same specificity in comparison with EB6, anti-CD4 (HP2.6), anti-DR (DI.12, IgG2a) In place of the commercially available mAbs JT3A, HP2.6, and DR1.12, of the same specificities can be used for example from Beckman coulter Inc., Fullerton, CA. EB6 and GL183 are commercially available from Beckman Coulter Inc., Fullerton, CA. XA-141 is not commercially available but EB6 can be used to control reconstitution of lysis as described in (Moretta et al., 1993).

Flow cytometry The cells were stained with the appropriate antibodies (30 mns at 4 ° C) followed by polyclonal anti-mouse antibodies conjugated with PE or FITC (Southern Biotechnology Associates Inc). The samples were analyzed by cytofluorometric analysis in a FACSAN apparatus (Becton Dickinson, Mountain View, CA).

Cytotoxicity experiments The cytolytic activity of the NK clones was assessed by a 51 Cr release assay in 4 standard hours. In which the effector NK cells were tested in Cw3 or Cw4 positive cell lines known for their sensitivity to NK cell lysis. All targets were used at 5000 cells per well in a microtiter plate and the Effector in the white ratio is indicated in the Figures (usually 4 effectors per target cells). Cytolytic analysis was performed with or without monoclonal antibody supernatant indicated at a medium dilution. The procedure is practically the same as that described in (Moretta et al., 1993).

Generation of novel mAb Mabs were generated by immunizing 5-week-old Balb C mice with activated polyclonal or monoclonal NK cell lines as described in (Moretta et al., 1990, the exhibition thereof is incorporated herein by reference). After different cell fusions, the mAbs were first selected for their ability to cross-react with the NK cell lines of EB6 and GL183 positive and the clones. The positive monoclonal antibodies were further selected for their ability for reconstitutive lysis by the NK EB6 positive or GL183 positive clones of the Cw4 or Cw3 positive targets respectively.

DF200, a novel monoclonal antibody against a common determinant of the human NK receptors of KIR2DL One of the monoclonal antibodies, the DF200 mAb, was found to react with various members of the KIR family including KIR2DL1, KIR2DL2 / 3. Regardless of NK cell staining with mAb DF200, both KIR2DL1 + and KIR2DL2 / 3 + were brilliantly stained (Figure 1). NK clones expressing one or the other (or both) of these HLA class I-specific inhibitory receptors were used as effector cells against target cells expressing one or more HLA-C alleles. As expected, KIR2DL1 + NK clones exhibited little, if any, cytolytic activity against target cells expressing the NK clones HLA-Cw4 and KIR2DL3 + that exhibit little or no activity on the Cw3 positive targets. However, in the presence of the mAb DF200 (used to mask their KIR2DL receptors) the NK clones became unstable to recognize their HLA-C ligands and exhibited significant cytolytic activity on the Cw3 or Cw4 targets. For example, the C1R cell line (CW4 + cell line EBV, ATCC No. CRL 1993) was not killed by NK KIR2DL1 + clones (CN5 / CN505), although the inhibition could be efficiently reversed by using either DF200 or a conventional anti-KIR2DL1 mAb. On the other hand, NK clones expressing the phenotype KIR2DL2 / 3 + KIR2DL1 (CN12) efficiently exterminated C1R and this extermination was unaffected by the mAb DF200 (Figure 2). Similar results were obtained with the NK KIR2DL2- or KIR2DL3- positive clones in the Cw3 positive targets.

Biacore analysis of the interactions mAb DF200 / KIR 2DL1 and mAb DF200 / KIR 2DL3 Materials and methods Production and purification of recombinant proteins. The recombinant proteins KIR2DL1 and KIR2DL3 were produced in E. coli cDNA according to the total extracellular domain of KIR2DL1 and KIR2DL3, amplified by PCR from vector 47.11 of clone pCDM8 (Biassoni et al, 1993; exposure thereof). incorporated herein by reference) and the ß-vector of clone RSVS (gpt) 183 (Wagtman et al, 1995, the disclosure thereof is incorporated herein by reference) respectively, using the following primers: Sense: 5 '-GGAATTCCAGGAGGAATTTAAAATGCATGAGGGAGTCCACAG-3' Anti-sense: 5 '-CCCAAGCTTGGGTTATGTGACAGAAAC__AGCAGTGG-3' They were cloned into the pML1 expression vector in the frame with a sequence encoding a biotinylation signal (Saulquin et al, 2003, the disclosure thereof being incorporated herein by reference). Protein expression was performed in the bacterial strain BL21 (DE3) (Invitrogen). The transfected bacteria were grown up to OD6_o = 0.6 at 37 ° C in medium supplemented with ampicillin (100 μg / ml) and incubated with 1 mm IPTG. The proteins were recovered from the inclusion bodies under denaturing conditions (8 M urea). The replication of the recombinant proteins was performed in Tris 20 mm, pH 7.8, L-arginine containing the buffer NaCl 150 mm (400 mm, Sigma) and β-mercaptoethanol (1 mm) at RT, when the concentration of urea in a six step dialysis (urea 4, 3, 2, 1, 0.5 and OM, respectively). Reduced and oxidized glutathione (sigma 5 mm and 0.5 mm respectively) was added during the dialysis steps with urea 0.5 and 0 M. Finally, the proteins were subjected to dialysis extensively against 10 mm Tris, pH 7.5, 150 mm NaCl buffer. The soluble refolded proteins were concentrated and then purified on a Superdex 200 size exclusion column (Pharmacia, AKTA system).

Biacore analysis. Measurements were made by surface plasmon resonance in a Biacore apparatus (Biacore). In all Biacore experiments the supplemented HBS buffer is 0.05% surfactant P20 served as the carrier buffer.

Protein immobilization. The recombinant K1R2DL1 and KIR2DL3 proteins were covalently immobilized with the carbonyl groups in the dextran layer on a CM5 detector chip (Biacore). The detector chip surface was activated with EDC / NHS (N-ethyl-N '- (3-dimethylaminopropyl) carbodiimidahydroxychloride and N-hydroxysuccinimide, Biacore). Proteins were injected into the coupling buffer (10 mm acetate pH 4.5). The deactivation of the remaining activated groups was carried out using 100 mm ethanolamine pH8 (Biacore).

Affinity measurements. For kinetic measurements, various concentrations of the soluble antibody (10 ~ 7 to 4 x 10"10 M) were applied to the immobilized sample, and measurements were made at 20 μl / min of continuous flow magnitude. Detector chip was regenerated using 5 μl of 10 mm NaOH injection -pH 11. The BIAlogue Kinetics Evaluation program (BIAevaluation 3.1, Biacore) was used for data analysis.

Results BIAcore analysis of mAb DF200 that binds immobilized KIR2DL1 and KIR2DL3.

KD: dissociation constant The soluble analyte (40 μl at various concentrations) was injected at a flow magnitude of 20 μl / min in HBS buffer, in dextran layers containing 500 or 540 reflectance units (RU), and 1000 or 700 RU of KIR2DL1 and KIR2DL3 respectively. The data are representative of 6 independent experiments.

Example 2: Improvement of ADCC by using a combination of Rituxan and an anti-KIR mAb Preparation of human NK clones. Blood mononuclear cells extracted from T lymphocytes by anti-CD3 immunomagnetic selection (Miltenyi) were plated under limiting dilution conditions, activated with phytohemagglutinin (PHA, for its acronym in English) (Biochrom KG, Berlin, Germany), and were cultured with interleukins (IL) -2 (Chiron B.V., Amsterdam, The Netherlands) and irradiated food cells. The cloning efficiencies were equivalent in all the donors and varied between 1 in 5 and 1 in 10 NK cells plated. The cloned NK cells were selected for alloreactivides by standard 51Cr release cytotoxicity against B lymphoblastoid cell lines transformed by Epstein-Barr virus of the HLA type known as an effector at the white ratio of 10: 1. Clones that exhibited a >lysis; 30% were marked as alloreactive. As a rule, the clones exhibit a lysis of either < 5% or > 40% Improvement of ADCC supplied by Rituxan medin and a NK KIR2DL1 positive cell clone The cytolytic activity of the NK clone was assessed by a standard 4 hour 51 Cr release assay, in which effector NK cells were tested on EBV Cw4 cell lines or Cw3 positive (CD20 positive), known for their sensitivity to NK cell lysis. All targets were used at 5000 cells per well in a microtiter plate and the effector (NK cell clone) at a white ratio is indicated in Figure 3. In certain experiments, the anti-CD20 rituximab was added (Rituxan, Idee) therapeutic chimeric at 5 μg / ml to the target effector mixture. In certain experiments, the EBß antibody (anti-KIR2DL1) was added at 10 μg / ml to the target effector mixture. This experiment showed that Rituxan alone does not essentially supply ADCC by the positive NK KIR2DL1 clone in the Cw4 positive target. The ADCC clone of KIR2DL1 was greatly improved in the presence of the anti-KIR2DL1 antibody.

Example 3 - Improvement of ADCC supplied by Campath by a positive NK KIR2DL1 cell clone In an experiment similar to that described in Example 2, Cw4 + PHA blasts were incubated in the presence of NK cells plus alumtuzumab (Campath, Berlex), the EBß antibody (at 100 μg / ml), or Campath and EBß. The results, shown in Figure 4, show that the presence of EBß exceptionally improves the ability of NK cells to decrease antigen cells: approximately 4% of target cells were lysed in the presence of Campath alone, while more of 30% of the cells were lysed in the presence of Campath plus EBß. All publications and patent applications mentioned in this specification are incorporated herein by reference in their entireties as if each individual publication or patent application had been specifically and individually indicated to be incorporated by reference. Although the above invention has been described in some detail by way of illustration and example for purposes of clarity and understanding, it will be readily apparent to one of ordinary skill in the art in light of the teachings of this invention that certain changes can be made and modifications to it without departing from the spirit or scope of the appended claims. Any combination of the elements described above in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or is clearly contradicted otherwise by context. The terms "a" and "an" and "the" and similar references in the sense in which they are used in the context to describe the invention should be construed to cover both the singular and the plural, unless indicated in the present in another way or clearly contradicted by context. The mention of variations of values herein is intended to simply serve as an abbreviated method to individually reference each separate value that falls within the variation, unless otherwise indicated herein, and each value separately is incorporated in the specification as if it were mentioned individually herein. Unless stated otherwise, all exact values provided herein are representative of the corresponding approximate values (eg, all exact example values provided with respect to the particular factor or measurement may also be considered to provide a measurement). corresponding approximate, modified by "approximately", when appropriate). All methods described herein may be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or example language (eg, "such as") provided in this, is simply intended to better understand the invention and has no limitation on the scope of the invention unless otherwise indicated. No language in the specification should be interpreted as indicating that any element is essential for the practice of the invention unless it is explicitly stated. The citation and incorporation of patent documents herein is for convenience only and does not reflect any view of the validity, patentability and / or possibility of application of these patent documents. The description herein of any aspect or mode of the invention using terms such as for example "comprising", "having", "including" or "containing" with reference to an element or elements is intended to provide support for a similar aspect or embodiment of the invention which "consists of", "consists essentially of", or "substantially comprises" that particular element or elements, unless otherwise stated or clearly contradicted by context (e.g., a composition described herein comprising a particular element should also be understood to describe a composition consisting of that element, unless otherwise stated or clearly contradicted by context). This invention includes all modifications and equivalents of the subject mentioned in the aspects or claims presented herein to the maximum extent permitted by applicable law.

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Claims (55)

1. NOVELTY OF THE INVENTION Having described the present invention, it is considered as a novelty and, therefore, the content of the following CLAIMS is claimed as property: 1. A method for the treatment of a disease in a human subject that needs it. , characterized in that it comprises: a) administering to the subject a compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell; and, b) administering to the subject a therapeutic antibody that can be bound by CDlβ.
2. 2. The method according to claim 1, characterized in that the therapeutic antibody has an Fe IgG1 or a human IgG3 portion.
3. The method according to claim 1 or 2, characterized in that the compound is an antibody or comprises a binding fragment with antigens thereof.
4. The method according to claim 1 to 3, characterized in that the therapeutic antibody is a monoclonal antibody or comprises a binding fragment with antigens thereof.
5. The method according to any of the preceding claims, characterized in that the therapeutic antibody is not conjugated with a radioactive or toxic entity.
6. 6. The method of any one of the preceding claims, characterized in that the compound inhibits an inhibitor receptor of an NK cell.
7. The method according to claims 1 to 5, characterized in that the compound stimulates an activating receptor of an NK cell.
8. The method according to any of the preceding claims, characterized in that the compound is a human, humanized or chimeric antibody or comprises a binding fragment with antigens thereof.
9. The method according to any of the preceding claims, characterized in that the therapeutic antibody is a human, chimeric humanized antibody or comprises a binding fragment with antigens thereof.
10. The method according to any of the preceding claims, characterized in that the therapeutic antibody is rituximab or campath.
11. The method according to claim 10, characterized in that the antibody is rituximab, and the antibody is administered at a dosage lower than 375 mg / m2 per week.
12. The method according to claim 10, characterized in that the antibody is iscampath, and the antibody is administered at a dosage lower than 90 mg per week.
13. The method according to any of the preceding claims, characterized in that the compound binds to at least one of the human receptors NKG2, KIR2DL or KIR3DL, and decreases the inhibition provided by NKG2, KIR2DL- or KIR3DL- of the cytotoxicity of NK cells .
14. The method according to any of the preceding claims, characterized in that the compound blocks an inhibitory receptor of an NK cell selected from the group consisting of KIR2DL1, KIR2DL2 / 3, KIR2DL4, KIR2DL5A, KIR2DL5B, KIR3DL1, KIR3DL2, KIR3DL3, LILRB1, NKG2A , NKG2C NKG2E and LILRB5.
15. The method according to any of the preceding claims, characterized in that the compound binds to a common determinant of the human KIR2DL receptors and decreases the inhibition provided by KIR2DL- of the cytotoxicity of NK cells.
16. The method according to claim 15, characterized in that the compound binds to a common determinant of the human receptors KIR2DL1, KIR2DL2, and KIR2DL3 and decreases the inhibition provided by KIR2DL1-, KIR2DL2-, and KIR2DL3- of the cytotoxicity of NK cells .
17. The method according to claim 16, characterized in that the compound inhibits the binding of a molecule of the HLA-C allele having a Lys residue at position 80 for a human KIR2DL1 receptor, and the binding of a molecule of the HLA-C allele. has an Asn residue at position 80 for the human KIR2DL2 and KIR2DL3 receptors.
18. 18. The method according to claim 13, characterized in that the compound binds to the same epitope as the monoclonal antibody DF200 produced by the hybridoma DF200, or the monoclonal antibody EBß.
19. 19. The method according to claim 13 to 17, characterized in that the compound competes with the monoclonal antibody DF200 produced by the hybridoma DF200, or the monoclonal antibody EBß, for binding to a KIR receptor on the surface of a human NK cell.
20. The method according to claim 13 to 17, characterized in that the compound is a monoclonal antibody DF200 produced by the hybridoma DF200 or a fragment or derivative thereof, or the monoclonal antibody EBß or a fragment or derivative thereof.
21. The method according to claim 7, characterized in that the compound is an antibody or comprises a binding fragment with antigens thereof.
22. The method according to claim 7 or 21, characterized in that the compound binds to a receptor selected from the group consisting of NKp30, NKp44, NKp4β, and NKG2D.
23. 23. The method according to claim 22, characterized in that the compound is derived or competes with a monoclonal antibody selected from the group consisting of AZ20, A76, Z25, Z231, and BAB281.
24. The method according to any of the preceding claims, characterized in that the therapeutic antibody and the compound are administered in the subject simultaneously.
25. 25. The method according to any of the preceding claims, characterized in that the compound is administered to the subject approximately one week after administration of the therapeutic antibody.
26. 26. The method according to any of the preceding claims, characterized in that the disease is a cancer, an infectious or immune disease.
27. The method according to any of the preceding claims, further characterized in that it comprises an additional step in which the activity or the number of NK cells in the patient is assessed before or after the administration of the compound.
28. The method according to claim 27, characterized in that it involves the additional steps: i) obtaining NK cells from a subject before administration; ii) incubating the NK cells in the presence of one more target cells that are recognized by the therapeutic antibody, in the presence or absence of the compound; and iii) assessing the effect of the compound on the ability of NK cells to decrease target cells; wherein a detection of this compound improves the ability of NK cells to decrease target cells, indicating that the compound is suitable for use in the method and the method is suitable for use with the subject.
29. 29. A pharmaceutical composition characterized in that it comprises: (a) a therapeutic antibody that can be bound by CD16, (b) a compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell, and (c) a pharmaceutically carrier acceptable.
30. 30. The composition according to claim 29, characterized in that the therapeutic antibody has a human or non-human primate Fe IgG1 or IgG3 portion.
31. 31. The composition according to the claims 29 or 30, characterized in that the compound is an antibody or comprises a binding fragment with antigens thereof.
32. 32. The composition according to claim 29 or 30, characterized in that the compound is a monoclonal antibody or comprises a binding fragment with antigens thereof.
33. 33. The composition according to claims 29 to 32, characterized in that the compound is a human, humanized or chimeric antibody or comprises a binding fragment with antigens thereof.
34. 34. The composition according to the claims 29 to 33, characterized in that the therapeutic antibody is a monoclonal antibody or comprises a binding fragment with antigens thereof.
35. 35. The composition according to claim 34, characterized in that the therapeutic antibody is a human, humanized or chimeric antibody or comprises a binding fragment with antigens thereof.
36. 36. The composition according to claims 34 or 35, characterized in that the antibody is not conjugated with a radioactive or toxic entity.
37. 37. The composition according to claims 29 to 36, characterized in that the compound inhibits an inhibitor receptor of an NK cell.
38. 38. The method according to claim 29 to 36, characterized in that the compound stimulates an activating receptor of an NK cell.
39. 39. The composition according to claims 29 to 38, characterized in that the therapeutic antibody is rituximab or CAMPATH.
40. 40. The composition according to the claims 29 to 37 and 39, characterized in that the compound blocks an inhibitor receptor of an NK cell selected from the group consisting of KIR2DL1, KIR2DL2 / 3, KIR2DL4, KIR2DL5A, KIR2DL5B, KIR3DL1, KIR3DL2, KIR3DL3, LILRB1, NKG2A, NKG2C NKG2E and LILRB5 .
41. 41. The composition according to claim 29, characterized in that the compound binds to a common determinant of the human KIR2DL receptors and decreases the inhibition provided by KIR2DL- of the cytotoxicity of NK cells.
42. 42. The composition according to claim 41, characterized in that the compound binds to a common determinant of the human receptors KIR2DL1, KIR2DL2, and KIRZDL3 and decreases the inhibition provided by KIR2DL1-, KIR2DL2-, and KIR2DL3- of the cytotoxicity of NK cells .
43. 43. The composition according to claim 41, characterized in that the compound inhibits the binding of a molecule of the HLA-C allele having a Lys residue at position 80 for a human KIR2DL1 receptor, and the binding of a molecule of the HLA-C allele. has an Asn residue at position 80 for the human KIR2DL2 and KIR2DL3 receptors.
44. 44. The composition according to claims 29 or 40 to 43, characterized in that the compound binds to the same epitope as the monoclonal antibody DF200 produced by the hybridoma DF200, the monoclonal antibody NKVSF1, or the monoclonal antibody EBβ.
45. 45. The composition according to claim 29 or 40 to 43, characterized in that the compound competes with the monoclonal antibody DF200 produced by the hybridoma DF200, the monoclonal antibody NKVSF1, or the monoclonal antibody EB6, to bind to a KIR receptor on the surface of a human NK cell.
46. 46. The composition according to claim 29 or 40 to 43, characterized in that the compound is a monoclonal antibody DF200 produced by the hybridoma DF200 or a fragment or derivative thereof, the monoclonal antibody NKVSF1 or a fragment or derivative thereof, or the antibody monoclonal EBß or a fragment or derivative thereof.
47. 47. The composition according to claim 38, characterized in that the compound binds to a receptor selected from the group consisting of NKp30, NKp44, NKp46, and NKG2D.
48. 48. A method for selecting a compound for administration together with a therapeutic antibody, the method characterized in that it comprises: i) providing a test compound that inhibits an inhibitory receptor or stimulates an activating receptor in NK cells; ii) incubating the therapeutic antibody with target cells specifically recognized by the therapeutic antibody in the presence of NK cells and in the presence or absence of the test compound; and iii) assessing the effect of the compound on the ability of NK cells to decrease target cells; wherein a detection of this compound improves the ability of NK cells to decrease target cells indicates that the compound is suitable for use in this method.
49. 49. The method according to claim 48, characterized in that the compound improves the ability of the therapeutic antibody to kill target cells by 30%.
50. 50. The method according to claim 48, characterized in that the compound improves the ability of the therapeutic antibody to destroy target cells by 50%.
51. 51. The method according to claim 48 to 50, characterized in that the compound is selected from the group consisting of an antibody, an antibody fragment, a monoclonal antibody, a fragment of a monoclonal antibody, a humanized antibody, a chimeric antibody, and a human antibody.
52. 52. The method according to claim 48 to 51, characterized in that the target cells are cancer cells, virally infected cells, or cells affected by an autoimmune disorder.
53. 53. The method according to claim 48 to 52, characterized in that the therapeutic antibody is rituximab or CAMPATH.
54. 54. A method for increasing the efficiency of a treatment including the administration of a therapeutic antibody that can bind by CDlβ in a subject, the method characterized in that it comprises administering to the subject before simultaneously with, or after administration of the therapeutic antibody , a therapeutically effective amount of a compound that blocks an inhibitory receptor or stimulates an activating receptor of an NK cell.
55. 55. The method according to claim 54, characterized in that the compound increases the efficiency of the treatment by improving ADCC in the subject.