MXPA06006327A - New class of gamma delta t cells activators and use thereof - Google Patents

Abstract

The present invention relates to a new class of compounds having&ggr;? T cells activating properties, a composition comprising these compounds and methods for regulating an immune response in a subject comprising the step of administering these compounds.

Description

NOVEDOSA CLASS OF GAMMA DELTA T-CELL ACTIVATORS AND USE THEREOF FIELD OF THE INVENTION The present invention relates to a novel class of compounds having T? D cell activating properties, a composition comprising these compounds and methods for regulating an immune response in an individual comprising the step of administering these compounds.

BACKGROUND OF THE INVENTION The majority of T? D cells of human peripheral blood express a? DTCR heterodimer encoded by V? 9 / Vd2 genes, some receptors of the NK lineage for MHC class I and almost do not express CD4 or CD8. It has been shown that these cells have cytolytic activity not restricted by strong MHC against virus-infected cells (Poccia et al (1999)), parasite-infected cells (Constant et al. (1995)), or tumor cells ( Fournie et Bonneville (1996)). These cells are also physiologically amplified in the context of several unrelated infectious diseases such as tuberculosis, malaria, tularemia, colibacillosis and also by B-cell tumors (for a review see Hayday, 2000). In addition to its anti-infective activity, short-term cytotoxicity tests have shown that V? 9 / Vd2 T cells can lyse a wide variety of tumor cell lines from a variety of sources: lymphoma and leukemia from lineages of B cell, T cell, or myeloid (Fisch et al., 1997; Selin et al., 1992; Sicard et al., 2001; Stur et al., 1990; Zheng et al., 2001a), breast tissue carcinoma (Bank et al., 1993), glioblastoma (Fujimiya et al., 1997; Yamaguchi et al., 1997), renal cell carcinoma (Choudhary et al., 1995; Kobayashi et al., 2001; Mitropoulos et al., 1994), nasopharyngeal carcinoma (Zheng et al., 2001b), lung adenocarcinoma, (Ferrarini et al., 1996). In microbes, V? 9 / Vd2 + lymphocytes spontaneously recognize a structurally related set of non-peptide antigens, known as natural phospho-antigens and alkylamines. In B-cell tumors, the nature of antigens for T? D cells remains unidentified. V? 9 / Vd2 + lymphocytes can also respond to a variety of virally infected, activated or tumor cell types without prior exposure. Again, in these situations, the responsible antigens remain unknown (for a review see Fisch, 2000). It has been shown that, in vitro, V? 9 / Vd2 2+ lymphocytes respond to synthetic drugs such as therapeutic aminobisphosphonates (reviewed in Espinosa, 2001), which lead to their activation in vitro. The recognition of natural non-peptidic antigens is mediated by? D TCR, through the amino acid residues located in both the V? 9- and Vd2 regions of CDR3. Although neither processing nor presentation by CD1 or MHC molecules is involved, activation of the V? 9 / Vd2 + lymphocyte by non-peptide antigens seems to require cell-to-cell contact (Lang, 1995; Morita, 1995; Miyagawa; , 2001, Rojas, 2002). It has been shown that the stimulating bacterial antigens are small non-peptidic compounds referred to classically as phospho-antigens (Behr et al., 1996, Belmant et al., 2000, Constant et al., 1994, Poquet et al., 1998, Tanaka et al. al., 1995), due to the presence of phosphate groups in most cases. V? 9 / Vd2 T cells can also be activated through endogenous metabolites (acting in the micromolar range) such as isopentenyl pyrophosphate or IPP (Espinosa et al., 2001b; Tanaka et al., 1995), which is produced through the conventional mevalonate route shared by both microorganisms and mammalian cells. The production of IPP in mammalian cells can be regulated in a positive way in situations of cell tension and transformation. In particular, a recent study has reported a correlation between the levels of endogenous production of IPP in tumor cells and their susceptibility to lysis mediated by V? 9 / Vd2 T cell (Gober et al., 2003). Also consistent with a direct contribution of the endogenous metabolites of the mevalonate pathway to the recognition of TV? 9 / Vd2 cell, the treatment of cells with pharmacological agents that prevent the biosynthesis of IPP (such as statins) or that lead to the accumulation of IPP (such as aminobisphosphonates, see below) lead respectively to the decrease or increase in the stimulatory properties of TV? 9 / Vd2 cells of the treated cells (Gober et al., 2003; Kato et al. , 2001). It is believed that aminobisphosphonates inhibit FPP synthetase, an enzyme in the mevalonate pathway, the inhibition of which causes the accumulation and release of upstream isoprenoid lipids such as IPP. The aminobisphosphonate compounds have been used in human therapy for the treatment of bone tissue metastases in cancer patients, and provide a first set of evidence for the in vivo expansion of V? 9 / Vd2 + human lymphocytes induced by phospho-agonists -antigen, which report increases in T? d cells in circulation within one to three weeks in human adults with multiple myeloma after intravenous therapeutic injection of 60-90 mg of pamidronate (Kunz ann et al., 1999). However, said compounds require that they be presented by cells that present antigen and can not produce substantial stimulation of V? 9 / Vd2 T cell activity as assessed by cytokine secretion in a pure V? 9 / Vd2 T cell culture. Likewise, pamidronate shows very low activation power of T? D cells, and it is reported that it achieves at most only an increase of twice as much in the T? D cell count (Wilhelm et al., 2003). Recently, a number of highly potent T? D cell activating pyrophosphate containing compounds have been described which directly activate T? D cells. In particular, the phosphalohydrin and phospho-epoxide compounds were described by the group of J.J. Fournie The compound (R, S) -3- (bromomethyl) -3-butanol-l-yl-diphosphate, also known as BrHPP (bromohydrin pyrophosphate) is currently used in clinical studies in humans that are performed to stimulate cell proliferation T? D ex vivo. Other pyrophosphate-containing compounds with high specific activity (EC50 in the nanomolar range or greater) are produced through an isoprenoid biosynthesis pathway called the "Rohmer" or "non-mevalonate" pathway, which is specific for prokaryotic microorganisms and eukaryotes (Feurle et al., 2002; Jomaa et al. (2003); Jomaa et al., 1999a; Jomaa et al., 1999b; Rohmer et al., 1993). Despite the foregoing, there remains a need for novel compounds that provide T? D cell activation, particularly compounds that have increased potency and / or preferred pharmacodynamic properties. Such compounds have particular advantages in non-life threatening indications or chronic therapeutic indications in which the therapies must be free of toxicity.

SUMMARY OF THE INVENTION The present invention now describes a novel class of compounds having T? D cell activating properties. This novel class of compounds comprises phosphorus-amidate esters. The inventors have discovered that the class of compounds described in the present invention have increased potency with respect to other compounds that modulate T? D cell activity previously evaluated by the inventors. In addition, the compounds can achieve a higher T? D cell activating effect in their CE100 (the efficient concentration of the composition that produces its maximum response or maximum effect with respect to said T? D activity) than other compounds. These compounds can be used to efficiently regulate the activity of T? D cells, in particular the activation and proliferation of T? D cells, preferably V? 9 / Vd2 T cells, in vivo, in an individual. These novel T? D cell activators can be used in accordance with any of the methods described in the present invention. These compounds are particularly suitable for immunotherapy, in particular for treating an individual having a tumor or an individual suffering from other diseases, in particular an infectious disease, an autoimmune disease or an allergic disease. The compounds according to the present invention can also be used as an adjuvant for vaccine. In one aspect, the invention provides a compound (T? D cell activator) of the formula (I): Formula (I) in which Cat + represents a cation or several cations, identical or different, organic or mineral (including proton); m is an integer from 1 to 3; B is 0, NH, or any group that can be hydrolyzed; Y = 0 ~ Cat +, an alkyl group of Cx-C3, an -AR group, or a radical that is selected from the group consisting of a nucleoside, an oligonucleotide, a nucleic acid, an amino acid, a peptide, a protein , a monosaccharide, an oligosaccharide, a polysaccharide, a fatty acid, a simple lipid, a complex lipid, a folic acid, a tetrahydrofolic acid, a phosphoric acid, an inositol, a vitamin, a co-enzyme, a flavonoid, an aldehyde , an epoxide and a halohydrin; A is 0, NH, CHF, CF2 or CH2; and R is a linear, branched, or cyclic, aromatic or non-aromatic, saturated or unsaturated C? -C5u hydrocarbon group, optionally interrupted by at least one heteroatom, wherein said hydrocarbon group comprises an algeryl, an alkylenyl, or an alkynyl, preferably an alkyl or an alkylene, which may be substituted with one or more substituents which are selected from the group consisting of: an alkyl, an alkylenyl, an alkynyl, an epoxyalkyl, an ary, a heterocycle , an alkoxy, an acyl, an alcohol, a carboxylic group (-COOH), an ester, an amine, an amino group (-NH2), an amide (-CONH2), an imine, a nitrile, a hydroxyl (-0H), an aldehyde group (-CHO), a halogen, a halogenoalguilo, a thiol (-SH), a thioalkyl, a sulfone, a sulfoxide, and a combination of them. In a preferred embodiment, said activator is a compound of the formula (X): wherein R3, R, and R5, identical or different, are a hydrogen or an alkyl group of C? ~ C3, W is -CH- or -N-, R5 is a C2-C3 acyl, an aldehyde, an alcohol of C? -C3, or a C2-C3 ester, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), B is O or NH, m is an integer of 1 to 3, and Y is 0 ~ Cat +, a nucleoside, or an -AR radical, in which A is O, NH, CHF, CF2 or CH2, and R is selected from the group consisting of 1), 2), or 3).

In a further aspect, said activator is a compound that is selected from the group consisting of: Formula (XI); Formula (XII) (XII) N-HDMAPP and formula (II) OH O O X- -C- - (CH2) n-N- -P-B- -P- Y H, H (ID Rl Ó Cat CfCat wherein in said formulas II, XI and XII: "X" is a halogen (which is preferably selected from I, Br and Cl), B is 0 or NH, m is an integer from 1 to 3, Rl is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), and n is an integer from 2 to 20, and Y is 0 ~ Cat +, a nucleoside, or an -AR radical, in which A is O, NH, CHF, CF2 or CH2 and R is selected from the group consisting of 1), 2), or 3). In additional modalities, the activator of T? d cell is a compound of the formula (III) In additional modalities, the activator of T? d cell is a compound of the formula (V) OH O O Br-C- - (CH2) 2-N P-O P-O, xCaA H2 H CH, 0 0 (V) N-BrHPP In additional modalities, the activator of T? d cell is a compound of the formula (VI): in which R1 is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different organic or mineral (including the proton), B is O or NH, m is an integer from 1 to 3, and n is a integer from 2 to 20, and Y is 0 ~ Cat +, a nucleoside, or an -AR radical, in which A is O, NH, CHF, CF2 or CH2 and R is selected from the group consisting of 1) , 2 or 3) . The present invention also provides a pharmaceutical composition comprising a T? D cell activator according to any of the embodiments described in the present invention. Methods are also provided for modulating, preferably activating, a T? D cell, the method comprising contacting a T? D cell with a T? D cell activating compound described in the present invention. As will be appreciated, the compounds of the invention can be used to activate T? D cell in vitro and in vivo. The T? D cell activated in vitro can be used in any appropriate method after activation, including in therapy or disease prevention. In a preferred example, activated T? D cells are administered to a mammal, preferably a human. In a preferred aspect, the invention encompasses a method of treatment comprising (a) putting a T? D cell in contact with a T? D cell activating compound described in the present invention and (b) administering the T? D cells of the step (a) to an individual. Methods for preparing T? D cells for such applications are known in the art, for example, they can be performed as described in US 10 / 505,252, filed on August 19 and 2004 PCT / FR 03/00585 presented on February 21, 2003, both for Romagne and Laplace, whose descriptions are incorporated in the present invention for reference. Methods for modulating, preferably activating a T? D cell comprising administering to an individual a T? D cell activator described in the present invention are also provided. In preferred embodiments, the inventions provide a method for treating or preventing a disease comprising administering to a subject a T cell activator described in the present invention in an amount sufficient to reduce or prevent said disease. The use of a T? D cell activator of the invention is also provided for the manufacture of a pharmaceutical composition for regulating T? D cells in a human individual. Preferably, said disease is a tumor or a proliferative disorder, an infectious disease, an autoimmune disease or an allergic disease. The invention also provides methods for the synthesis of phosphoramidate compounds. In one aspect, the invention provides a method for preparing a diphosphoramidate mono-ester compound comprising: (a) reacting an alkyl halide R-X in a coupling step with a reagent of diethylphosphoramidate or diethylchlorophosphate type; (b) reacting the compound prepared in step (a) in a saponification step with which the O-ethyl groups are removed; and (c) reacting the compound prepared in step (b) in a phosphorylation step with which a mono-ester of diphosphoramidate is prepared, wherein R is a linear, branched, or cyclic, aromatic or non-aromatic, saturated or unsaturated C1-C50 hydrocarbon group, optionally interrupted by at least one heteroatom, wherein said hydrocarbon group comprises an alkyl, an alkylenyl , or an alkynyl, preferably an alkyl or an alkylene, which may be substituted with one or more substituents which are selected from the group consisting of: an alkyl, an alkylenyl, an alkynyl, an epoxyalkyl, an aryl, an heterocycle, an alkoxy, an acyl, an alcohol, a carboxylic group (-COOH), an ester, an amine, an amino group (-NH2), an amide (-CONH2), an imine, a nitrile, a hydroxyl (- OH), an aldehyde group (-CHO), a halogen, a halogenoalkyl, a thiol (-SH), a thioalkyl, a sulfone, a sulfoxide, and a combination thereof, and wherein X is a portion that can be be displaced by a diethylphosphoramidate group under appropriate conditions. Depending on the type and reactivity of the functional groups provided by R, the person skilled in the art can adapt the following examples, if necessary, including the protection / deprotection phases of the sensitive functional groups or those that can interact with the reaction of copulation. In an X-modality it is an NH2 group and said R-X compound is reacted in a coupling step with a diethylchlorophosphate compound. In another embodiment, X is selected from the group consisting of I, Br, and Cl. In another aspect, the invention provides a method for preparing a compound (E) -2- (4-azido-2-methylbut-2-enyloxy) tetrahydro-2J7-pyran, which comprises providing a compound (E) -2- ( 4-chloro-2-methylbut-2-enyloxy) tetrahydro-2i? -pyran and reacting said compound with a sodium azide in a biphasic mixture of water-pentane in the presence of phase transfer catalyst. The modalities and additional details are provided later in the present invention.

BRIEF DESCRIPTION OF THE FIGURES Figure 1 shows an in vitro dose-response curve and the EC5u values for the compound of the invention N-HDMAPP and the reference compounds BrHPP and HDMAPP. The compound of the invention demonstrates not only a potency increased by 3-4 times with respect to the following more potent compounds, but also an increased absolute activation of T? D cells as observed by release of TNFa.

DETAILED DESCRIPTION OF THE INVENTION Definitions Within the context of the present invention, the term "regulating the activity of T? D cells" designates causing or favoring an increase in the number and / or biological activity of said cells in an individual. Regulate, therefore, includes without limitation, modulate (e.g., stimulate) the expansion of said cells in an individual and / or, for example, induce cytokine secretion (e.g., TNFa or IFNα). As indicated, T? D cells typically represent between about 1-10% of the total circulating lymphocytes in a healthy adult human subject. The present invention can be used to significantly increase the population of T? D cells in an individual, in particular to reach at least 10%, 12%, 15%, 20%, or 30-90% of total circulating lymphocytes , typically 40-90%, more preferably from 50 to 90%. In typical embodiments, the invention allows the selective expansion of T? D cells in an individual, to reach 60-90% of the total lymphocytes in circulation, preferably 70-90%, more preferred from 80 to 90%. Regular also includes, in addition or alternatively, modulating the biological activity of the T? D cells in an individual, in particular their cytolytic activity or their cytokine secretion activity. The invention defines novel conditions and strategies for increasing the biological activity of T? D cells towards target cells. In cases where "comprising" is used, it may be replaced by "consisting essentially of", more preferred with "consisting of". In cases where numerical terms are used previously and subsequently in the present invention, they are intended to include the numbers representing the upper and lower limits. For example, "between 1 and 3" represents an interval "from and including 1 to and including 3", and "in the interval from 1 to 3" can mean "from and including 1 to and including 3". The same is true in cases where words that designate numbers (for example, "three") are used instead of numbers (for example "3"). Where "approximately" is used in connection with a number, this preferably means the number +/- 15%, more preferably the number plus 5%, even more preferred the number itself without "approximately", for example , "approximately 100" can mean "from and including 85 up to and including 115". Where "approximately" is used in connection with numerical ranges, for example "1 approximately to approximately 3", or "between approximately 1 and approximately 3", preferably the definition of "approximately" given for a number in the last sentence is applies to each number that defines the start and end of a separate interval. Preferably, where "approximately" is used in connection with any numerical values, the "approximately" may be suppressed. "Weekly" means "approximately once a week" (which means that more than one treatment is carried out with an interval of approximately one week between treatments), the word "approximately" in this case means preferably +/- 1 day ( that is to say, that is translated in "every 6 to 8 days"); more preferably, "weekly" means "once every 7 days". As used in the present invention, the term "EC50" with respect to regulating the activity of T? D cells, refers to the efficient concentration of the present compositions that produce 50% of their response or maximum effect with respect to said T cell activity? d. As used in the present invention, the term "CEioo" with respect to regulating T? D cell activity, refers to the effective concentration of the present compositions that produce their response or maximum effect with respect to said cell activity. T? D.

Novel class of T? D lymphocyte activators: phosphoramidate esters The novel class of compounds described by the inventors of the present invention comprise phosphoramidate esters. The inventors have discovered that compounds of this class show an increase in potency with respect to other compounds that modulate T? D cell activity previously evaluated by the inventors. In addition, the compounds of the invention can achieve a greater T? D cell activating effect in their CEioo (the efficient concentration of the composition that produces its response or maximum effect with respect to said activity of T? D cells) than other compounds. Although not wishing to be limited to the theory, the inventors of the present invention propose that the presence of an NH group may result in a modified bond - generally increased binding strength - to the objective of the compound, probably due to binding considerations of H compared, for example, with the pyrophosphate ester compounds. This modification in H binding considerations may provide different pharmacological properties, for example, target binding affinity, ADME properties (absorption, distribution, metabolism and excretion). In further preferred embodiments, the compounds of the invention also desirably have in vivo stability properties, preferably longer half-lives than other available compounds. The increased stability in blood may be useful to achieve enhanced global in vivo T? D cell stimulation. The novel class of T? D lymphocyte activators according to the present invention comprises the compounds of the formula (I): Formula (I) in which Cat + represents a cation or several cations, identical or different, organic or mineral (including proton); m is an integer from 1 to 3; B is O, NH, or any group that can be hydrolyzed; Y = 0"Cat +, an alkyl group of C? -C3, an -AR group, or a radical that is selected from the group consisting of a nucleoside, an oligonucleotide, a nucleic acid, an amino acid, a peptide, a protein, a monosaccharide, an oligosaccharide, a polysaccharide, a fatty acid, a single lipid, a complex lipid, a folic acid, a tetrahydrofolic acid, a phosphoric acid, an inositol, a vitamin, a co-enzyme, a flavonoid, a aldehyde, an epoxide and a halohydrin, A is 0, NH, CHF, CF2 or CH2, and R is a C, -C50 hydrocarbon, linear, branched, or cyclic, aromatic or non-aromatic, saturated or unsaturated, optionally interrupted by at least one heteroatom, wherein said hydrocarbon group comprises an alkyl, an alkylenyl, or an alkynyl, preferably an alkyl or an alkylene, which may be substituted with one or more substituents which are selected from the group consisting of of: an alkyl, an alkylenyl, an alkyne ilo, an epoxyalkyl, an aryl, a heterocycle, an alkoxy, an acyl, an alcohol, a carboxylic group (-C00H), an ester, an amine, an amino group (-NH2), an amide (-C0NH2), an imine, a nitrile, a hydroxyl (-0H), an aldehyde group (-CH0), a halogen, a halogenoalkyl, a thiol (-SH), a thioalkyl, a sulfone, a sulfoxide, and a combination of them.

In a particular embodiment, substituents as defined above are substituted by at least one of the substituents specified above. Preferably, the substituents are selected from the group consisting of: a Ci-Ce alkyl, a C2-C6 alkylenyl, a C2-C6 alkynyl, a C2-C5 epoxyalkyl, an aryl, a heterocycle, a Ci-Ce alkoxy, a C2-C6 acyl, a C1-C6 alcohol, a carboxylic group (-COOH), a C2-C5 ester, a C? -C6 amine, an amino group (-NH2) , an amide (-C0NH2), an imine of C? -C6, a nitrile, a hydroxyl (-OH), an aldehyde group (-CHO), a halogen, a halogenalkyl of C? -C6, a thiol (-SH), a thioalkyl of C? -C6, a sulfone of C? -C6, a sulfoxide of CI-CT, and a combination thereof. More preferably, the substituents are selected from the group consisting of: a C 1 -C 6 alkyl, a C 2 -C 6 epoxyalkyl, a C 2 -C 6 alkylenyl, a C 6 -C 6 alkoxy, an acyl C2-Cd, a Ci-Ce alcohol, a C2-C6 ester, a Ci-Cβ amine, a C-C6 imine, a hydroxyl, an aldehyde group, a halogen, a Ci-Ce halogenalkyl, and a combination of them. Even more preferably, the substituents are selected from the group consisting of: a C3-C6 epoxyalkyl, a C3-C3 alkoxy, a C2-C3 acyl, a C3-C3 alcohol, an ester of C2-C3, an amine of C? -C3, an imine of C? -C3, a hydroxyl, a halogen, a halogenalguile of C? -C3, and a combination thereof, and a combination thereof. Preferably, R is a C3-C25 hydrocarbon group, more preferred is a C5-C10 hydrocarbon group. In the context of the present invention, the term "alkyl" means more specifically a group such as methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, hexadecyl, heptadecyl, octadecyl, nonadecyl, eicosyl, heneicosyl, docosyl and the other isomeric forms thereof. Alkyl of C? -C6 more specifically means methyl, ethyl, propyl, isopropyl, butyl, isobutyl, tert-butyl, pentyl, hexyl and the other isomeric forms thereof. Alkyl of Ci-C3 more specifically means methyl, ethyl, propyl or isopropyl. The term "alkenyl" refers to an alkyl group defined above in the present invention having at least one unsaturated ethylene bond and the term "alkynyl" refers to an alkyl group defined above that has at least one acetylene-type bond unsaturated C2-C6 alkylene includes an ethenyl, a propenyl (1-propenyl or 2-propenyl), a 1-methylpropenyl or 2-methylpropenyl, a butenyl (1-butenyl, 2-butenyl, or 3-butenyl), a methylbutenyl, a 2-ethylpropenyl, a pentenyl (1-pentenyl, 2-pentenyl, 3-pentenyl, 4-pentenyl), a hexenyl (1-hexenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 5-hexenyl), and the other isomeric forms thereof. C2-C6 alkynyl includes ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 1-pentynyl, 2-pentynyl, 3-pentynyl, 4-pentynyl, 1-hexynyl, 2- Hexynyl, 3-Hexynyl, 4-Hexynyl or 5-Hexynyl and the other isomeric forms thereof. The term "epoxyalkyl" refers to an alkyl group defined above in the present invention having an epoxide group. More particularly, C2-C6 epoxyalkyl includes epoxyethyl, epoxypropyl, epoxybutyl, epoxypentyl, epoxyhexyl, and the other isomeric forms thereof. Epoxyalkyl of C2-C3 includes epoxyethyl and epoxypropyl. The "aryl" groups are mono-cyclic, bi-cyclic or tri-cyclic aromatic hydrocarbons having from 6 to 18 carbon atoms. Examples include a phenyl, α-naphthyl, β-naphthyl or anthracenyl group, in particular. The "heterocycle" groups are groups containing 5 to 18 rings comprising one or more heteroatoms, preferably 1 to 5 endocyclic heteroatoms. These may be monocyclic, bi-cyclic or tri-cyclic. These can be aromatic or non-aromatic. Preferably, and more specifically for R5, these are aromatic heterocycles. Examples of aromatic heterocycles include pyridine, pyridazine, pyrimidine, pyrazine, furan, thiophene, pyrrole, oxazole, thiazole, isothiazole, imidazole, pyrazole, oxadiazole, triazole, thiadiazole and triazine groups. Examples of bicycles include in particular quinoline, isoquinoline and quinazoline groups (for two 6-membered rings) and indazole, benzimidazole, benzoxazole, benzothiazole and indazole (for a 6-membered ring and a 5 membered ring). Non-aromatic heterocycles comprise in particular piperazine, piperidine, etc. "Alkoxy" groups correspond to the alkyl groups defined above in the present invention attached to the molecule by an -O- (ether) bond. Ci-Cβ alkoxy includes methoxy, ethoxy, propyloxy, butyloxy, pentyloxy, hexyloxy and the other isomeric forms thereof. C 1 -C 3 alkoxy includes methoxy, ethoxy, propyloxy, and isopropyloxy. "Acyl" groups correspond to the alkyl groups defined above in the present invention bound to the molecule by a -CO- (carbonyl) group. Acyl of C2-C6 includes acetyl, propylacyl, butylacyl, pentylacyl, hexylacyl, and the other isomeric forms thereof. Acyl of C2-C3 includes acetyl, propylacyl and isopropylacyl. Groups "alcohol" corresponds to the alkyl groups defined above in the present invention containing at least one hydroxyl group. Alcohol can be primary, secondary or tertiary. "C" alcohol includes methanol, ethanol, propanol, butanol, pentanol, hexanol and the other isomeric forms thereof. C? ~ C3 alcohol includes methanol, ethanol, propanol and isopropanol. "Ester" groups correspond to the alkyl groups defined above in the present invention linked to the molecule by a -COO- (ester) bond. Ester of C2-C6 includes methyl ester, ethyl ester, propyl ester, butyl ester, pentyl ester and the other isomeric forms thereof. Ester of C2-C3 includes methyl ester and ethyl ester. "Amine" groups correspond to the alkyl groups defined above in the present invention attached to the molecule by an -N- (amine) bond. C? -C6 amine includes methylamine, ethylamine, propylamine, butylamine, pentylamine, hexylamine and the other isomeric forms thereof. C? -C3 amine includes methylamine, ethylamine, and propylamine.

"Imine" groups correspond to the algilo groups defined above in the present invention having a bond (-CN = N-). Imine of Ci-Ce includes methylimine, ethylimine, propylimine, butylimine, pentylimine, hexylimine, and the other isomeric forms thereof. Imine of C1-C3 includes methylimine, ethylimine and propylimine. The halogen may be Cl, Br, I or F, more preferably Br or F. "Haloalogyl" groups correspond to the alkyl groups defined above in the present invention having at least one halogen. The groups may be monohalogenated or polyhalogenated containing the same or different halogen atoms. For example, the group may be a trifluoroalkyl (CF3-R). Halogenoalkyl of Ci-Cg includes halogenomethyl, halogenoethyl, halogenopropyl, halogenobutyl, halogenopentyl, halogenohexyl and the other isomeric forms thereof. Halogenoalkyl of C1-C3 includes halogenomethyl, halogenoethyl, and halogenopropyl. "Thioalkyl" groups correspond to the alkyl groups defined above in the present invention attached to the molecule via a -S- (thioether) linkage. Thioalkyl of C? -C6 includes thiomethyl, thioethyl, thiopropyl, thiobutyl, thiopentyl, thiohexyl and the other isomeric forms thereof. C1-C3 thioalkyl includes thiomethyl, thioethyl, and thiopropyl.

"Sulfone" groups correspond to the algilo groups defined above in the present invention linked to the molecule by a -S00- (sulfone) bond. Sulfone of C? -C6 includes methylsulfone, ethylsulfone, propylsulfone, butylsulfone, pentylsulfone, hexylsulfone and the other isomeric forms thereof. Sulfone of C? -C3 includes methylsulfone, ethylsulfone and propylsulfone. "Sulfoxide" groups correspond to the alkyl groups defined above in the present invention bound to the molecule by an -SO- (sulfoxide) group. Ci-Cß sulfoxide includes methyl sulfoxide, ethyl sulfoxide, propyl sulfoxide, butyl sulfoxide, pentyl sulfoxide, hexyl sulfoxide and the other isomeric forms thereof. C? ~ C sulfoxide includes methyl sulfoxide, ethyl sulfoxide, propyl sulfoxide and isopropyl sulfoxide. "Heteroatomo" means N, S, or O. "Nucleoside" includes adenosine, thymine, uridine, cytidine, and guanosine. In a particular embodiment, the hydrocarbon group is a cycloalguilenyl such as a cyclopentadiene or a phenyl, or a heterocycle such as a furan, a pyrrole, a thiophene, a thiazole, an imidazole, a triazole, a pyridine, a pyrimidine, a pyran , or a pyrazine. Preferably, the cycloalkylenyl or the heterocycle is selected from the group consisting of a cyclopentadiene, a pyrrole or an imidazole. In a preferred embodiment, the cycloalkylenyl or the heterocycle is substituted with an alcohol. Preferably, said alcohol is a C1-C3 alcohol. In another embodiment, the hydrocarbon group is an alkylenyl with one or more double bonds. Preferably, the alkylenyl group has a double bond. Preferably, the alkylenyl group is an alkylene group of C3-C? Or, more preferably, an alkylenyl group of C-C. Preferably, said alkylenyl group is substituted by at least one functional group. More preferred, the functional group is selected from the group consisting of a hydroxy, a C1-C3 alkoxy, an aldehyde, a C2-C3 acyl, or a C2-C3 ester. In a more preferred embodiment, the hydrocarbon group is butenyl substituted with a -CH2OH group. Optionally, said alkenyl group can be the trans (E) or cis (Z) isoform, more preferably, a trans (E) isoform. In a more preferred embodiment, the alkylenyl group is the (E) -4-hydroxy-3-methyl-2-butenyl compound. In another preferred embodiment, the alkylenyl group is an isopentenyl, a dimethylallyl or a hydroxydimethylallyl. In a further embodiment, the hydrocarbon group is an alkyl group substituted with an acyl. More preferred, the hydrocarbon group is an alkenyl group of C4-C7 substituted with an acyl of C? ~ C3. In a further preferred embodiment, R is selected from the group consisting of: 1) wherein n is an integer from 2 to 20, Ri is an alkyl group of C? ~ C3, and R2 is a halogenated C? -C3 alkyl, a (C1-C3) alkoxy-C1-C3 alkyl, a halogenated C 2 -C 3 acyl or a C 2 -C 3 alkoxy (C 2 -C 3) -acyl. Preferably, Ri is a methyl or ethyl group, and R2 is a halogenated methyl (-CH2-X, where X is a halogen), a halogenated C2-C3 acetyl, or (C1-C3) alkoxy-acetyl. The methyl or halogenated acetyl can be monohalogenated, dihalogenated or trihalogenated. Preferably, n is an integer from 2 to 10, or from 2 to 5. In a more preferred embodiment, n is 2. In a most preferred embodiment, n is 2, R is a methyl and R2 is a halogenated methyl, more preferred is a monohalogenated methyl, even more preferred a methyl bromide. In a particularly preferred embodiment, n is 2, Rx is a methyl, R2 is a methyl bromide. In a more preferred embodiment, R is 3- (bromomethyl) -3-butanol-1-yl. wherein n is an integer from 2 to 20, and Ri is a methyl or ethyl group. Preferably, n is an integer from 2 to 10, or from 2 to 5. In a more preferred embodiment, n is 2 and Rx is a methyl. 3) wherein R3, R and R5, identical or different, are a hydrogen or a C1-C3 alkyl group, W is -CH- or -N-, and Re is an acyl of C2-C3, an aldehyde, a C1-C3 alcohol, or a C2-C3 ester. More preferably, R5 is a methyl and R3 and R are a hydrogen. More preferably, Rβ is -CH2-0H, -CHO, -CO-CH3 or -C0-0CH3. Even more preferred, R6 is -CH2-0H. Most preferably, W is -CH-. Optionally, the double bond between W and C is in a trans (E) or cis (Z) conformation. More preferred, the double bond between W and C is in the trans (E) conformation. Group Y can allow the design of a prodrug. Therefore, Y is an enzyme-labile group that can be cut in particular regions of the individual. Group Y can also be a group for target selection. In a preferred embodiment, Y is 0"Cat +, an -AR group, or a radical that is selected from the group consisting of a nucleoside, a monosaccharide, an epoxide and a halohydrin. enzymes, preferably, Y is 0 ~ Cat +, an -AR group, or a nucleoside In a first preferred embodiment, Y is 0"Cat +. In a second preferred embodiment, Y is a nucleoside. In a preferred embodiment, Cat + is H +, Na +, NH +, K +, Li +, (CH 3 CH 2) 3 NH +, lysine, or any other suitable pharmaceutically acceptable cation. In a preferred embodiment, A is O, CHF, CF2 or CH2 More preferred, A is O or CH2. In a preferred embodiment, B is O or NH. More preferred, B is O. In a preferred embodiment, m is 1 or 2. More preferred, m is 1. In a particular embodiment, the phosphoramidate esters according to the present invention comprise the compounds of the formula (II): wherein X is a halogen (which is preferably selected from I, Br and Cl), B is 0 or NH, m is an integer from 1 to 3, Rl is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), and n is an integer from 2 to 20, and Y is 0 ~ Cat +, a nucleoside, or an -AR radical, in which A is O , NH, CHF, CF2 or CH2 and R is selected from the group of 1), 2) or 3). Preferably, Y is 0"Cat +, or a nucleoside, most preferred, Y is 0 ~ Cat +, preferably Rx is a methyl, preferably n is 2. Preferably, X is a bromide. O. Preferably, m is 1 or 2. More preferred, m is 1. For example, the phosphoramidate esters according to the present invention comprise the compounds of the formula (III): in which X, Rl, n, m and Y have the aforementioned meaning. In a preferred embodiment, the phosphoramidate esters according to the present invention comprise the compounds of the formula (IV): (IV) wherein X is a halogen (which is preferably selected from I, Br and Cl), Rl is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton) ), and n is an integer from 2 to 20. Preferably, R 1 is a methyl. Preferably, n is 2. Preferably, X is a bromide. In a more preferred embodiment, the phosphoramidate esters according to the present invention comprise the compound of the formula (V): (V) N-BrHPP Preferably xCat + is 1 or 2 Na +. In a particular embodiment, the phosphoramidate esters according to the present invention comprise the compounds of the formula (VI): in which R1 is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), B is 0 or NH, m is an integer from 1 to 3, and n is an integer from 2 to 20, and Y is 0 ~ Cat +, a nucleoside, or an -AR radical, in which A is 0, NH, CHF, CF2 or CH2, and R is selected from the group of 1) , 2 or 3) . Preferably, Y is 0"Cat +, or a nucleoside, most preferred, Y is 0 ~ Cat +, preferably R1 is a methyl, preferably n is 2. Preferably, B is 0. Preferably, m is 1 or 2. More preferred, m is 1. For example, the phosphoramidate esters according to the present invention comprise the compounds of the formula (VII): in which Rl, n, m and Y have the meaning mentioned above.

In a preferred embodiment, the phosphoramidate esters according to the present invention comprise the compounds of the formula (VIII): wherein R1 is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), and n is an integer from 2 to 20. Preferably, R1 is a methyl. Preferably, n is 2. In a more preferred embodiment, the phosphoramidate esters according to the present invention comprise the compound of the formula (IX): (IX) N-EpoxPP Preferably xCat + is 1 or 2 Na +. In a particular embodiment, the phosphoramidate esters according to the present invention comprise the compounds of the formula (X): (X) wherein R3, R4, and R5, identical or different, are a hydrogen or C? -C3 alkyl group, W is -CH- or -N-, R6 is an acyl of C-C3, an aldehyde, an alcohol of C1-C3, or a C2-C3 ester, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), B is O or NH, m is an integer from 1 to 3, and Y is 0"Cat +, a nucleoside, or an -AR radical, in which A is O, NH, CHF, CF2 or CH2, and R is selected from the group of 1), 2) or 3). preference, Y is 0 ~ Cat +, or a nucleoside.Most preferred, Y is 0 ~ Cat +, More preferred, R3 and R4 are a hydrogen and R5 is a methyl.Most preferred, R6 is -CH2-0H, -CHO, C0 -NH2, -NH2, or -CO-OCH3 Even more preferred, R6 is -CH2-0H Most preferred, W is -CH- Preferably, B is O. Preferably, m is 1 or 2. More preferred , m is 1. Optionally, the double bond between W and C is in trans (E) or cis (Z) conformation.Most preferred, the double bond between W and C is in co Transformation (E) For example, the phosphoramidate esters according to the present invention comprise the compounds of the formula (XI): in which R3, R4, R5, R6, W,, and Y have the meaning mentioned above. Preferably, W is -CH- or -N-. Preferably, R3 and R4 are hydrogen. Preferably, R5 is a methyl. Preferably, R6 is -CH2-0H. In a more preferred embodiment, the phosphoramidate esters according to the present invention comprise the compound of the formula (XII): (XII) N-HDMAPP These compounds can be produced in accordance with various techniques, for example using methods described in PCT publications nos. WO 00/12516, WO 00/12519, WO 03/050128, and WO 03/009855, the descriptions of which are incorporated in the present invention for reference. In a more preferred embodiment, the synthetic T? D lymphocyte activating compound is selected from the group consisting of N-HDMAPP, N-Epox-PP, and N-BrHPP, more preferably N-HDMAPP and N-BrHPP , even more preferably N-HDMAPP. The phosphoramidate esters according to the present invention can be prepared, for example, by the following reactions (Reactions A, A (1), A (2), A (3), B, C or C (1).

REACTION SCHEME FOR MONOESTERS DIFOSFORAMIDATE Reaction A O o ALTERNATIVE REACTION SCHEME FOR MONOESTERS DIFOSFORAMIDATE Reaction A (l) O O CCLCN / CH, CN H | | || R-NH2 R-N P-O P-OH > 2 eq. of Bu4NH2P04 | | OH OH The diphosphoramidate monoesters in accordance with the present invention can also be prepared using the following two reactions (Reactions A (2), A (3)). These synthesis schemes are preferred for larger-scale preparations of diphosphoramidate monoesters and involve the formation of a monophosphoramidate monoester intermediate which is of interest for pharmaceutical development because it is considered a potential metabolite or degradation product.

ALTERNATIVE REACTION SCHEME FOR MONOESTERS OF DIFOSFORAMIDATE Reaction A (2) Reaction A (2) can be conveniently used when the starting alkyl precursor (R-NH 2) can not be easily obtained, the starting molecule being preferably an alkyl halide RX, in which X = I, Br or Cl. a) TMSBr TMSBr / purifier L¡NH-P? (OEt) 2 O of acid O (prepared insitu) H II b) Bu4NF or Bu4N0H H || R- X - * - R- N - P - OEt R- N - P - O? U4N + Coupling step I Saponification step I OEt 0"Bu4N + Morpholinophosphate / pyridine Q Q CCl3CN / CH3C / < 2eq. (B? UU44"N) H2P04 H _ * + Phosphorylation step i r 0" Cat + 0"Cat + The coupling step of A (2) involves the in situ preparation of a lithium salt from the deprotonation of commercially available diethyl phosphoramidate (NH2-P (OEt) 2). This coupling step can be carried out following the procedure reported by Cox et al (2002), whose description is incorporated in the present invention for reference. The saponification step of reaction A (2) involves a two-step procedure which leads to a complete removal of the 0-ethyl ester functional groups. This reaction should preferably be carried out under neutral or basic conditions in order to avoid hydrolysis of the phosphoramidate bond (P-NH bond). This reaction can be carried out with trimethylsilyl bromide (TMSBr) with subsequent removal of the resulting TMS esters with tetrabutylammonium fluoride (Bu4NF) as described in Valentijn et al (1991) or using TMSBr in the presence of Sim-collidine (acid scavenger) as described in Valentijn (1995). ) with subsequent basic hydrolysis of the resulting TMS esters with tetrabutylammonium hydroxide (Bu4NOH). The descriptions of the above references are incorporated in the present invention for reference. The phosphorylation step of reaction A (2) can be carried out in two ways: (i) reaction of the intermediate monophosphoramidate with a morpholinophosphate reagent (tetrabutylammonium salt), whose preparation from commercially available di-methyl chlorophosphate is presented in more detail below (Reaction A (2) (a)). This reaction can be carried out following the procedure of Valentijn et al. (1991) as applied for the synthesis of pyrophosphonate analogues; or (ü) reaction of the intermediate monophosphoramidate with trichloroacetonitrile (CC13CN) as a coupling reagent and less than 2 equivalents of the commercially available tetrabutylammonium (Bu4N) H2P04 dihydrogenphosphate.

PREPARATION OF A REAGENT OF MORPHOLINOPHOSPHATE Reaction A (2) (a) ALTERNATIVE REACTION SCHEME FOR MONOESTERS OF DIFOSFORAMIDATE Reaction (3) a) TMSBr TMSBr / debugger TEA / C1-P0 (0H) 2 9 of acid ° (0 ° C) H I I b) Bu4NF Ó Bu4N0H H || - NH, * - R - N - P - OEt * • R - N - P - 0"Bu4? Coupling step I Saponification step I OEt 0"Bu4N + Morpholinophosphate / pyridine or O O Ca 3 CN / CH 3 CN / < 2eq. of (Bu4) H2P04 H I I | | Phosphorylation step * "R- N-POP-OCa 1 + I + O'Cat O'Caf The coupling step of reaction (3) involves the reaction of an alkylamine precursor with diethyl chlorophosphate commercially available in the presence of triethylamine ( TEA) .This reaction can be carried out following the procedure described in Nikolaides et al., (Conversion of Amines to Phosphoesters: decyl diethyl phosphate, Organic Syntheses, CV 9,194). The conditions for the saponification and phosphorylation steps are similar to those reported previously (Reaction A (2)). Depending on the type and reactivity of the functional groups provided by R, one skilled in the art can adapt the synthesis examples presented in the present invention, including, if necessary, the protection / deprotection phases of the sensitive functional groups or of those that can interact with the copulation reaction.

REACTION SCHEME FOR MONOESTERES OF IMIDO- DIFOSFORAMIDATO Reaction B 0 0 EDC or EDC / NHS R - NH R- N H I PI- -N "I PI - OH Na4HN06P2 (pH <6.5) OH OH REACTION SCHEME FOR TRIFOSFORAMIDATE MONOESTERS Reaction C O O O ALTERNATIVE REACTION SCHEME FOR MONOESTERS OF TRIFOSFORAMIDATE Reaction C O O or CC CN / CH3CN H II R - NH2 »• R - N P - O P - O P - OH > 3 eq. of Bu4NH2P04 j | | OH OH OH Reactions A, B and C can be carried out as described in Sato et al (1990) and Chu et al (1983) using l-ethyl-3- (3-dimethylaminopropyl) -carbodi-imide (EDC) as a coupling reagent. Inorganic reagents Na2HP207 (disodium pyrophosphate), NaHN06P2 (tetrasodium imidodiphosphate) and Na5O? 0P3 (pentasodium triphosphate) can be obtained commercially. The N-hydroxysuccinimide compound (NHS) is often used to aid in the coupling of carbodiimide in the presence of EDC (Seghal &Vijay, 1994). The descriptions of the above references are incorporated in the present invention for reference.

Reactions A, B and C can also be carried out in non-aqueous solvents with carbodiimide reagents such as DCC (N, Nf-dicyclohexylcarbodiimide) using organic diphosphate or triphosphate salts. Carbodi-imides have been widely used in the synthesis of ortho-phosphate and pyrophosphate esters, nucleotides, cyclic phosphates, oligo-ribonucleotides, polynucleotides, nucleoside-5'-phosphoroamidates, and mixed anhydrides (Azzi et al., 1984), whose description is incorporated in the present invention for reference. The alternative reactions A (l) and C can be carried out following the procedure described in Zhang and Poulter (1993), the description of which is incorporated in the present invention for reference, with trichloroacetonitrile (CC13CN) as a coupling reagent. The inorganic reagent tetrabutylammonium dihydrogen phosphate (Bu4N) H2P04 can be obtained commercially. The above reactions involve the protection of the sensitive functional groups of the R-NH2 compound or that react with the carbodiimide (EDC) or trichloroacetonitrile (CC13CN) reagents. The phosphoramidate monoesters can be purified by preparative C18 HPLC in accordance with the method reported by Zhang and Poulter (1993). Or by preparative silica gel chromatography using the ammonia isopropanol eluents in accordance with the methods of International Patent Publication No. WO 03/050128 filed December 5, 2002. The descriptions of the above references are incorporated in the present invention. for reference. The compounds comprising a nucleoside such as the group Y can be prepared, for example, by the following reactions. Depending on the type and reactivity of the functional groups provided by Y, the person skilled in the art can adapt the following examples, including, if necessary, the protection / deprotection phases of the sensitive functional groups or those that can interact with the reaction of copulation.

Nucí- O- V - NH - PP - »- R- NH - PPO - Nucí acetonitrüo Reaction D O Nucí- O- V R- NH - PPP - * - R - NH - PPPO - Nucí acetonrtrüo Reaction E in which -OV is a good leaving group starting with choosing V, for example, from tosyl, mesyl, triflyl, brosyl or bromine, PP represents the pyrophosphate group, PPP represents the triphosphate group, R- has the aforementioned meaning and Nucí is a nucleoside. Preferably, Nucl-OV is selected from the group consisting of: 5'-O-tosyladenosine, 5'-O-tosyluridine, 5'-O-tosylcytidine, 5'-O-tosylthymidine or 5'-O-tosyl -2 '-deoxy-adenosine. For example, for the compound with R of group 1), the reaction procedure may be as follows: Cr? 2 H2 II Nuci - O V II R, C (CH2) n - NHPP Rj C (CH2) n - NHPPO - Nucy Acetonitplo CH2 OH I I X2, H20 I R, C (CH2) n - NHPPO - uci * - X - H2C - C n (H2C) - HPPO - Nucí Rl in which -O-V is a good leaving group starting with choosing V, for example, from tosyl, mesyl, triflyl, brosyl or bromine, PP represents the pyrophosphate group and Nuci is a nucleoside. Preferably, Nucl-OV is selected from the group consisting of: 5 '-O-tosyladenosine, 5'-O-tosyluridine, 5', - O-tosylcytidine, 5'-O-tosylthymidine or 5 '-O- tosyl-2'-deoxyadenosine as described in Davisson et al, (1987), the disclosure of which is incorporated in the present invention for reference. The nucleophilic substitution reaction can be carried out under conditions similar to those described by Davisson et al, (1987); and Davisson et al. (1986), whose descriptions are incorporated in the present invention for reference. This reaction can also be used to prepare the compound comprising a monosaccharide as a group Y. In this case, Nucl-O-V is replaced by MonoSac-0-V, in which Monosac is monosaccharide. For example, it is possible to use the MonoSac-OY group corresponding to the compound Methyl-6-O-tosyl-alpha-D-galactopyranoside as described in the publication by Nilsson and Mosbach, (1980), incorporated in the present invention for reference , or the man-made triflate compound which can be obtained commercially. This reaction can also be used to prepare a compound comprising an oligosaccharide as a group Y. In this case, Nucl-O-V is replaced by oligoSac-O-V, in which oligoSac is an oligosaccharide. For example, it is possible to use the oligoSac-OY group corresponding to the compound 6A-0-p-toluenesulfonyl-β-cyclodextrin as described in the publication (Organic syntheses, Vol. 77, p 225-228, the description of which is incorporated in the present invention for reference). This reaction can be used to prepare a compound comprising a polysaccharide as a group Y. In this case, Nucl-O-V is replaced by polySac-O-V, in which polySac is a polysaccharide. For example, it is possible to use the polySac-0-Y group which corresponds to the tosylated polysaccharide as described in the publication by Nilsson et al., (1981); and Nilsson and Mosbach, (1980), whose descriptions are incorporated in the present invention for reference. This coupling technique based on activation of the hydroxyl groups of a polysaccharide support by tosylation allows covalent coupling in an aqueous medium or in an organic medium. This reaction can also be used to prepare a compound comprising an aldehyde derivative as a group and choosing, instead of Nuci, a derivative including a protected aldehyde functional group in the form of an acetal or any other group that protects this functional group. Alternatively, compounds comprising a nucleoside as a group Y can be prepared by the following reaction: l) Nucl-0-PPP, carbodiimide DMF / Methanol R-? H2 ** R-? HPPPO -? ucl 2) Triethylamine, DMF Reaction F in which PPP represents the triphosphate group, R- has the As mentioned above, DMF is dimethylformamide, and ? ucl is a nucleoside. This reaction can be carried out in conditions similar to those described by Knorre et to the. (1976), or by Bloom et al., Patent E.U.A. ?or. ,639,653 (1997), whose descriptions are incorporated in the present invention for reference, from alcohol and a nucleotide with the formula Nucl-O-PPP. For example, for the compound with R of group 1), the reaction procedure may be as follows: CH2 l) Nucl -? - PPP, carbodiimide CH2 | | DMF / Methanol | | R, C (CH2) n - NH2 »- R, C (CH2) p - NHPPPO-Nucl 1 2 n 2 2) TrietflaminaIDMF X 2 U CH2 OH I I Xa, H20 I R, C (CH,) n - NHPPPO-Nucl - * - X - CH2 - C n (H2C) - NHPPPO - Nucl 1 Jn pHneutro? go, in which PPP represents the triphosphate group, DMF is dimethylformamide, and Nuci is a nucleoside. This reaction can also be applied to the preparation of oligonucleotide 5'-triphosphate esters as indicated by the authors of the Knorre et al. (1976). Compounds that comprise a nucleic acid such as group Y, more particularly a ribonucleic acid, can be prepared under conditions similar to those described in the publication of F. Huang et al (1997). The authors describe a universal method from catalytic RNA that can be applied to any molecule comprising a free terminal phosphate group. Compounds structurally related to the phosphohalohydrin group such as isopentenyl pyrophosphate or thiamine pyrophosphate are used or mentioned by these authors (see page 8968 of F. Huang et al (1997)). It should also be mentioned that the experimental conditions for the coupling procedure (in particular pH conditions) described in the section "Reaction of Isolate 6 pppRNA with phosphate containing Nucleophiles" on page 8965 are compatible with the presence of a halohydrin functional group. Compounds comprising an amino acid, a peptide or a protein derivative as a group Y can be obtained using the well-known reactivity of its primary amine or thiol functional group on an epoxide functional group (SN2 reaction). This type of coupling classically involves an intermediate group still called a "linker" having an epoxide functional group. An example of a reaction procedure using this type of coupling is provided below.

R'- SH R- A- NHPPO - CH2 - CH2OH - S - R 'Y And Reaction G wherein PP represents the pyrophosphate group, R- has the above-mentioned meaning and R '-SH is an amino acid, a peptide or a protein derivative. The first phase can be carried out under conditions similar to those described by Davisson et al. (1987) and Davisson et al, (1986), the disclosures of which are incorporated herein by reference, from the tetrabutylammonium salt of the starting compound and commercially available compounds such as glycidyl tosylate or epichlorohydrin. This reaction can also be carried out with triphosphate compounds. Alternatively, a primary amine R '-NH2 can be used in place of R'-SH. Without the reaction with R'-SH, the first reaction can be used to prepare a compound comprising an epoxide derivative. Alternatively, compounds comprising an amino acid, a peptide or a protein derivative as a group Y can be prepared by the following reaction: 1) R'-NH2, carbodiimide DMF / Methanol R-NH-PPP * - R- NH- PPO- P- NH-R '2) Triethylamine, DMF Reaction H in which PPP represents the triphosphate group, PP represents the pyrophosphate group, P represents the group phosphate, R- has the above-mentioned meaning and R '-NH is an amino acid, a peptide or a protein derivative. The reaction can be carried out under conditions similar to those described by Knorre et al. (1976), which description is incorporated in the present invention for reference, from the compound (R-NH-PPP) and an amino acid, peptide or a protein with the formula R'-NH2. This reaction involves the protection of the sensitive functional groups of the compound R'-NH2 or that can react with the carbodiimide (in particular, the carboxyl functional group). Tri or tetra-n-butylammonium salts of phosphoric, pyrophosphoric, triphosphoric, tetraphosphoric or polyphosphoric acid can be prepared from the corresponding commercially available acids. Derivatives with a related structure such as the methan-triphosphonic acid derivatives described in the publication by Liu et al (1999), the disclosure of which is incorporated herein by reference, may also be prepared in accordance with the reaction procedure. The aforementioned reactions can be extrapolated to a very large spectrum of molecules or biomolecules using the reactivity of the hydroxyl, amine, phosphate or thiol functional groups. In this way, inositol derivatives can be prepared in accordance with reactions D or E by activation of the hydroxyl functional group. Folic acid derivatives (vitamin B9) or tetrahydrofolic acid can be prepared according to the G or H reactions by reacting the reactivity of the primary amine functional group. Of course, other types of coupling may be considered and a person skilled in the art may have access to a large choice of reactions. Therefore, coupling can be used by phosphorylation of carboxylic acid or phenolic groups for the formation of fatty acids, lipid or some fiavonoid derivatives. The T? D lymphocyte activating phosphoramidate ester compound can be a molecule that is produced ex vivo or in vitro. This may be an endogenous ligand purified or artificially produced in some other way (for example, by chemical synthesis, or by microbiological processes) or a fragment or derivative thereof, or an antibody having substantially the same antigenic specificity. The phosphoramidate esters according to the present invention can preferably selectively activate the V? 9Vd2 T lymphocytes. The selective activation of V? 9Vd2 T lymphocytes indicates that the compound has a selective action towards specific populations of cells, and essentially does not activate other subtypes of T cells, such as Vdl T cells. Said selectivity, as described in the present application, suggests that preferred compounds may cause a selective or targeted activation of the proliferation or biological activity of V? 9Vd2 T lymphocytes. In a preferred aspect, the T? D cell activator can increase the biological activity of T? D cells, preferably increasing the activation of T? D cells, increasing in particular the secretion of cytokine from T? Do cells increasing the cytolytic activity of T? d cells, without stimulating or also stimulating the expansion of T? d cells. In typical embodiments, a T? D cell activator allows the secretion of cytokine by T? D cells to increase by at least 2, 3, 4, 10, 50, 100 times, as determined in vitro. Cytokine secretion and cytolytic activity can be evaluated using any appropriate in vitro test, or those provided in the examples of the present invention. For example, cytokine secretion can be determined with the methods described in Espinosa et al. (J. Biol. Chem., 2001, Vol. 276, exemplary 21, 18337-18344), which describe the measurement of TNF-α release in a biological test using TNF-? Sensitive cells. In brief, 104 T? D cells / cavity with stimulus plus 25 units of IL2 / cavity are incubated in 100 μl of culture medium for 24 hours at 37 ° C. Then, 50 μl of supernatant is added to 50 μl of WEHI cells seeded at a density of 3 x 104 cells / well in culture medium plus actinomycin D (2 μg / ml) and LiCl (40 mM) and incubated for 20 hours at 37 ° C. the viability of cells sensitive to TNF-α is measured with a bromide test 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium. 50 μl of 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide (Sigma) is added.; 2.5 mg / ml in phosphate-buffered saline solution) per well, and after 4 hours of incubation at 37 ° C, 50 μl of buffer solution is added for solubilization (20% SDS, 66% dimethylformamide, pH 4.7), and the absorbance (570 nm) is measured. The levels of TNF-α release are then calculated from a standard curve obtained using purified human rTNF-a (PeproTech, Inc., Rocky Hill, NJ). Interferon-? released by activated T cells using an enzyme-linked immunosorbent assay in sandwich. 5 × 10 4 T d d cells / cavity with stimulus plus 25 units of IL2 / well are incubated in 100 μl of culture medium for 24 hours at 37 ° C. Then, 50 μl of supernatant is collected for the enzyme-linked immunosorbent assay using mouse monoclonal antibodies (BIOSOURCE, Camarillo, CA). A preferred test for cytolytic activity is a 51 Cr release test. In the example tests, the cytolytic activity of the Tαd cells is measured against normal and tumor autologous target cell lines, or control sensitive target cell lines such as the Daudi line and the control resistant target cell line such as Raj i in a 4-hour 51Cr release test. In a specific example, target cells are used in amounts of 2 x 10 3 cells / well and labeled with 100 μCi of 51 Cr for 60 minutes. The Effector / Objective (E / T) ratio varies from 30: 1 to 3.75: 1. Specific lysis (expressed as a percentage) is calculated using the standard formula [(experimental release - spontaneous release! spontaneous release tota) l0?] Use of T? D lymphocyte activators in accordance with the present invention The invention relates to a pharmaceutical composition comprising a T? D cell activator in accordance with the present invention. More particularly, said pharmaceutical composition comprises a therapeutically effective amount of T? D cell activator, optionally together with a pharmaceutically acceptable carrier. The invention also encompasses the use of a T? D activator in accordance with the present invention for the manufacture of a pharmaceutical preparation, preferably for the treatment of a cancer, an infectious disease, an autoimmune disease or an allergic disease. In one aspect, the invention describes a method for regulating T? D cells in a human subject, said method comprising the step of administering, at least in one treatment, a therapeutically effective amount of a T? D cell activator in accordance with the present invention, optionally together with a pharmaceutically acceptable carrier. More specifically, said method aims to stimulate T? D cells in a human individual. In a particular embodiment, the amount of said T? D cell activator is sufficient to expand the population of T? D cells in an individual to reach at least 10%, 15%, 20%, 30%, 40%, 50 % or 60%, or between 30-90% of total circulating lymphocytes. In another embodiment, the amount of said T? D cell activator is sufficient to induce an increase of at least 10-fold in the population of T? D cells in an individual. Preferably, said population of T? D cells is evaluated between day 4 and day 8 after administration of said T? D cell activator, most preferably on day 5, 6 or 7 after administration of T? D cells. said T cell activator? d. Preferably, said population of T? D cells is evaluated by flow cytometry. Preferably, said T? D cells are V? 9 / Vd2 cells. In a preferred embodiment, the invention relates to a method for treating a cancer, an infectious disease, an autoimmune disease or an allergic disease in an individual, said method comprising the step of administering, at least in one treatment, a Therapeutically effective amount of a T? d cell activator according to the present invention, optionally together with a pharmaceutically acceptable carrier. In the above methods and uses, the individual of preference is a human subject, such as an individual having a cancer, an infectious disease, an autoimmune disease or an allergic disease. Indeed, the invention is suitable for treating all conditions caused by or associated with the presence of pathological cells that are sensitive to T? D cell lysis. The invention is particularly suitable for stimulating the anti-tumor immunity of an individual having a solid or hematopoietic tumor. Preferably, said tumor is selected from the group consisting of tumors of the lung, colorectal, prostate, breast tissue or epidermoid head or neck. In a preferred aspect of the invention, said tumor is a renal cancer, preferably a metastatic renal cancer. Alternatively, said tumor is selected from the group consisting of a melanoma, ovarian cancer, pancreatic cancer, neuroblastoma, head or neck cancer, bladder cancer, kidney cancer, brain cancer and gastric cancer. In preferred embodiments, the compounds can be used for the treatment of cancer as described in the international patent application no. PCT / IB2003 / 006375, filed December 2, 2003, the description of which is incorporated in the present invention. The invention is also suitable for stimulating an anti-viral immune response in an individual having an infection caused by a virus that is selected from HIV, CMV, EBV, influenza virus, HCV, HBV, etc. The compounds of the invention are also suitable in methods for stimulating an immune response in an individual having an infection caused by a pathogen causing tuberculosis, malaria, tularemia, colibacillosis, etc. The compounds of the invention are also suitable in methods of treating (e.g., to stimulate an immune response in) an individual having an autoimmune disease, such as diabetes, multiple sclerosis, rheumatoid arthritis, etc. or an individual who has an allergic disease, including asthma, hyper-responsiveness of airways, etc. In preferred embodiments the compounds are used in therapeutic indications and in accordance with the teachings of the international patent application no. WO2000US0026684 filed on September 28, 2000 by Gelfand, Born, Lahn, and Kanehiro; international patent application no. WO 00/00182, filed June 24, 1999 by Jomaa; and provisional patent application E.U.A no. 60 / 564,959 filed on April 26, 2004 by Tiollier, the descriptions of each of the references are incorporated in the present invention for reference. Preferably, the dose (individual administration) of a phosphoramidate ester compound according to the present invention for treatment is between about 1 μg / kg and about 1.2 g / kg. It will be appreciated that the above dosages refer to a group of compounds, and that each particular compound may vary at the optimum doses, as described below in the present invention for the example compounds. However, the compounds are preferably administered in a sufficient dose to significantly increase the biological activity of T? D cells or to significantly increase the population of T? D cells in an individual. Said preferred dose is administered to the human by intravenous (iv) administration for 2 to 180 minutes, preferably 2 to 120 minutes, more preferably for about 5 minutes to about 60 minutes, or even more preferably for about 30 minutes or 60 minutes approximately. In the preferred exemplary compounds, a compound of formula II to XII is administered in a dose (individual administration) between about 1 μg / kg and about 1.2 g / kg, preferably between about 10 μg / kg and 1.2 g / kg approximately, more preferably between about 20 μg / kg and about 100 mg / kg. More preferably, the dose (individual administration) for tri-weekly or tetra-weekly treatment (treatment every three weeks or every third week) is between approximately 1 μg / kg and approximately 1.2 g / kg, preferably between approximately 10 μg / kg and 20 mg / kg approximately, more preferably between about 10 μg / kg and about 100 mg / kg. This preferred dose is administered to the human by intravenous (iv) administration for 2 to 180 minutes, preferably 2 to 120 minutes, more preferably for about 5 minutes to about 60 minutes, or even more preferably for about 30 minutes or 60 minutes approximately. The active ingredients can be administered through different routes, typically by injection or oral administration. The injection can be carried out in various tissues, such as intravenously, intraperitoneally, intra-arterially, intra-muscularly, intra-dermally, subcutaneously, etc. The preferred routes of administration for the activators are intravenous and intra-muscular. The preferred routes of administration for the cytokine are subcutaneous, intravenous and intra-muscular. The invention provides a method for regulating T? D cell activity in a mammalian subject, the method comprising administering to an individual in need thereof an effective amount of a T? D cell activator in accordance with a treatment cycle in which allows the T? d cell activity, preferably the T? d cell rate (number of T? d cells), to return to the substantially basal number before a second administration of the compound. As described later in the present invention, in preferred embodiments, at least about one week, but more preferably at least two weeks, is required for the number of cells T? D of a patient returns to the substantially basal number.

Shorter cycles of approximately 7 days do not allow for the appropriate stimulation of T cell activity. The course of a preferred cycle is a cycle of at least one week, but more preferably at least one bi-weekly cycle (at least 14 days approximately), or more preferably at least three weeks, although cycles are preferred at any point between bi-weekly and four-weekly. They are also effective and are scheduled cycles of up to 8 weeks, for example 5 weeks, 6 weeks, 7 weeks or 8 weeks. In a preferred embodiment, the administration of the T? D cell activator occurs on the first day of a bi-weekly to quad-weekly cycle (i.e., a repetitive cycle of 14 to 28 days). In a preferred embodiment, the T? D cell activator is administered only on the first day of the bi-weekly to quadri-weekly, or preferably tri-weekly cycle. As mentioned, an individual is preferably treated for at least two cycles, or more preferably for at least three cycles. In another aspect, the treatment may continue for a greater number of cycles, for example, at least 4, 5, 6 or more cycles may be contemplated. Optionally, T? D cell activators can also be used in combination with a cytokine.

Preferably, said cytokine is interleukin 2 (IL-2) (Proleukin ™, Chiron, Emeryville CA, USA) or any biologically active fragment, variant or analogue thereof, i.e., any fragment, variant or analog that can be bound to an IL-2 receptor and that can induce T? d cell activation in the method of this invention. Preferably, said T? D activator and interleukin-2 polypeptide are administered separately to the individual. Therefore, the methods of the invention also comprise administering a cytokine. Although the compounds of the invention can be used with or without additional administration, a cytokine can be administered in a preferred aspect, wherein said cytokine can increase the expansion of a T? D cell population treated with a T cell activating compound. ? d, preferably in which the cytokine can induce an expansion of a T? d cell population that is greater than the expansion resulting from the administration of the T? d cell activating compound in the absence of said cytokine. A preferred cytokine is an interleukin-2 polypeptide. A cytokine having T-cell proliferation-inducing activity, most preferred the interleukin-2 polypeptide, is administered at low doses, typically over a time interval of between 1 and 10 days. The T? D cell activator is preferably administered in a single dose, and typically at the start of a cycle. Preferably, the interleukin-2 polypeptide is administered at a daily dose of between 0.2 and 2 MU per day, even more preferably between 0.2 and 1.5 MU, even more preferred between 0.2 and 1 MU. The daily dose of cytokine, preferably an interleukin-2 polypeptide, is administered as a single injection or in two injections. In preferred aspects, a cytokine, most preferred IL-2, is administered daily for a period of up to about 10 days, preferably for a period between about 3 and 10 days, or more preferably for about 7"days. administration of the cytokine begins on the same day (for example within 24 hours of) as the administration of the T cell activator d.For example, in one aspect, the cytokine is administered every day, while in other aspects it is not necessary The cytokine is administered every day When the cytokine is administered for about 7 to about 14 days, a four-weekly treatment cycle is preferred When the first component is administered for about 4 days, a 3-day treatment cycle is preferred per week In preferred embodiments, the compounds may be used in accordance with any of the methods described in the international application. Patent No. PCT / IB2003 / 006375, filed December 2, 2003, the disclosure of which is incorporated in the present invention for reference. The above methods and treatments can be used alone or in combination with other active agents or treatments. For example, for the treatment of tumors, the invention can be used in combination with other anti-tumor agents or treatments, such as chemotherapy, radiotherapy or gene therapy. The invention also relates to a product comprising a T? D cell activator according to the present invention and an interleukin-2 polypeptide, for separate use, for regulating the activity of T? D cells in a mammalian subject. The invention relates to a vaccine composition comprising a T? D cell activator in accordance with the present invention. The invention also relates to the use of a T? D cell activator in accordance with the present invention as a vaccine adjuvant. Accordingly, the present invention describes methods and compositions for increasing and / or increasing the immune response against an antigen in a mammal, specifically a human, which involves the joint immunization of the mammal with (i) a composition comprising an antigen y ( ii) an adjuvant comprising a phosphoramidate ester compound according to the present invention. Preferably, said composition comprises an antigen comprising a pathogen, microorganism or parasite annihilated, inactivated or attenuated. In another aspect, said composition comprises an antigen, which preferably comprises a polypeptide, lipid, polysaccharide, glycoprotein, glycolipid or enriched or purified nucleic acid antigen. Preferably, said composition comprises at least 1, 2, 3, 4, 5, 10 or 15 different antigens, for example at least 1, 2, 3, 4, 5, 10 or 15 polypeptides, or nucleic acids encoding for said polypeptides, different. In preferred embodiments, the compounds can be used as described in the provisional patent application E.U.A. do not. 60/564, 959, filed April 26, 2004, the disclosure of which is incorporated in the present invention for reference. The adjuvant composition may comprise an effective amount of a phosphoramidate ester compound according to the present invention, said amount being an effective amount that allows the induction of a humoral response, induction of a cytotoxic T lymphocyte (CTL) response, or the induction of both a humoral response and a CTL response of the adjuvant composition with respect to said at least one antigen. Preferably the phosphoramidate ester compound according to the present invention is present in an amount effective to produce a greater immunological effect to induce a humoral response, a cytotoxic T lymphocyte (CTL) response or both a humoral response and a response of CTL when co-administered with an antigen that the immunological effect that occurs when said antigen is administered in the absence of adjuvant. The antigen component of the composition can be selected from virtually any antigen, antigenic determinant or hapten of medical or veterinary interest, and in particular for those antigens for which an increase in immunogenicity is desired. Therefore, the present invention relates to the use of a phosphoramidate ester compound according to the present invention, more preferably N-HDMAPP or N-BrHPP, as an adjuvant for vaccine. The present invention also relates to a vaccine composition comprising an antigen or a combination of antigens, and a phosphoramidate ester compound according to the present invention, more preferably N-HDMAPP or N-BrHPP. Preferably, said composition comprises a therapeutically effective amount of antigen and an amount that increases an immune response or enhances an immune response, of the T? D cell activ of phosphoramidate ester. Preferably, said vaccine composition prevents a microbial infection. Said microbial infection is caused by a microbe that is selected from the group consisting of viruses, fungi, parasites, yeasts, bacteria, and protozoa. In a particular embodiment, said vaccine composition is a BCG vaccine composition. Alternatively, said vaccine composition prevents or is a treatment against a tumor. The present invention also relates to a vaccine case comprising an appropriate container containing a vaccine composition according to the present invention, more particularly comprising an antigen or a combination of antigens, and a phosphoramidate ester compound of according to the present invention, more preferably N-HDMAPP or N-BrHPP. Optionally, said vaccine may comprise two separate suitable containers, one of which contains the antigen or the combination of antigens and the other contains a phosphoramidate ester compound according to the present invention, more preferably N-HDMAPP or N-BrHPP. Optionally, said container can be a syringe. Alternatively, said vaccine case comprises one or two containers and a syringe. The present invention relates to a method for improving the potency of a vaccine in an individual, or to immunize an individual against a disease, more particularly a microbial infection, comprising the steps of: administering to said individual a composition that comprises an antigen or a combination of antigens; and jointly administering to said individual a phosphoramidate ester compound according to the present invention, more preferably N-HDMAPP or N-BrHPP, more particularly an amount thereof to increase an immune response. Preferably, the activator cell T? D when coadministered with a composition comprising an antigen, is administered in an amount sufficient to increase an immune response over that observed with said composition comprising an antigen in the absence of activator of cell T? d. Preferably, said composition comprising an antigen comprises a pathogen, microorganism or parasite annihilated, inactivated or attenuated. In another aspect, said composition comprising an antigen preferably comprises a polypeptide, lipid, polysaccharide, glycoprotein, glycolipid or nucleic acid antigen. The present invention also relates to a method of immunizing an individual against a disease, more particularly a microbial infection in an individual comprising administering to said subject (i) a composition comprising an antigen, and (ii) a phosphoramidate ester compound according to the present invention, more preferably N-HDMAPP or N-BrHPP. Preferably, the T? D cell activator is administered in an amount that increases an immune response. Preferably, the T? D cell activator and the composition comprising an antigen are administered as an individual vaccine composition in a therapeutically effective amount. Preferably, said T? D cell activator is together with a pharmaceutically acceptable carrier. In a first aspect, said administrations of said antigen or combination of antigens and said T? D cell activator are simultaneously. In a second aspect, said administrations of said antigen or combination of antigens and said T? D cell activator are sequentially. More particularly, said T? D cell activator can be administered before, concurrent with or subsequent to the administration of an antigen or a combination of antigens to an individual for immunization purposes. Preferably, said antigen or combination of antigens are microbial antigens, preferably viral, bacterial, fungal, protozoan, yeast or parasite antigens. In a preferred embodiment, said antigen is an antigen of Mycobacterium bovis. Optionally, said antigen or combination of antigens is a tumor antigen. The additional aspects and advantages of this invention are described in the following examples, which should be considered as illustrative and not as limitative of the field of this application.

EXAMPLES EXAMPLE 1 Production of (E) -4-hydroxy-3-methylbut-2-enyl (N-HDMAPP) pyrophosphoramidate Preparation of (E) -4-chloro-2-methylbut-2-en-l-ol 16 ml (179 mmol) of nitrogen are added under nitrogen.

TiCl4 to 360 mL of CH2C12. The solution is cooled to 90 ° C and a solution of 10.0 g (119 mmol) of 2-methyl-2-vinyloxirane in 50 ml of CH2C12 is added dropwise, keeping the temperature below -80 ° C. The red solution is then stirred at 80 ° C for 2 hours and quenched with 600 ml of 1M HCl. The organic phase is separated and the aqueous phase is extracted with 3 x 500 ml of Et20. The combined organic phases are dried with MgSO 4, filtered and evaporated at 350 mbar at 25 ° C to obtain 12.02 g (99.7 mmol, 84% yield) of 4-chloro-2-methylbut-2-en-1-ol as a brown oil. The raw product is used directly in the next step.

Preparation of (E) -2- (4-chloro-2-methylbut-2-enyloxy) tetrahydro-2H-pyran To a solution of 11.5 g (95.37 mol) of 4-chloro-2-methylbut-2-en-1 -ol in 120 ml of CH2C12 is added 26 ml (286.11 mmoles) of dihydropyran (DHP). The solution is cooled to 0 ° C and 2.4 g (9.53 mmoles) of pyridinium p-toluenesulfonate (PPTS) are added in portions. The solution is stirred for 3 hours at 0 ° C. The organic phase is washed with 3 x 50 ml of water, dried with Na 2 SO, filtered and concentrated to obtain the crude product. The product is then purified by chromatography on silica gel using heptane / EtOAc (9/1) as eluent. 12.35 g (60.33 immoles, 64% yield) of the protected allylic alcohol are isolated as a colorless oil.

Preparation of (E) -2- (4-azido-2-methylbut-2-yloxy) tetrahydro-2H-pyran A solution of 11.43 g (175.86 mmoles) of sodium azide (NaN3) and 2.20 g (6.45 g) is added. mmoles) of tetrabutylammonium hydrogensulfate in 60 ml of water to a solution of 12.0 g (58.62 mmol) of (E) -2- (4-chloro-2-methylbut-2-enyloxy) tetrahydro-2H-pyran in 300 ml of pentane The reaction mixture is stirred at room temperature overnight. The organic phase is separated and the aqueous phase is extracted with 3 x 150 ml of Et20. The combined organic phases are washed with brine, dried with Na2SO4, filter and concentrate to obtain 11.21 g (53.06 mmoles, 90% yield) of a mixture of azide isomers. No purification is carried out because unfeasible silica gel separation is observed. The product is used without further purification for the next step.

Preparation of (E) -3-methyl-4- (tetrahydro-2H-pyran-2-yloxy) but-2-en-l-amine 26 g (98.8 mmol) of triphenylphosphine are added (PPh3) to a solution of 11.0 g (52.07 mmol) of the crude azide obtained in the previous step and 18 ml of water in 180 ml of THF. The solution is stirred at room temperature overnight and the solvent is evaporated. The resulting crude product is purified by chromatography with silica gel (eluent: CH2Cl2 / MeOH / Et3N 9/1 / 0.5) to obtain 6.47 g (34.91 mmol, 67% yield) of purified allylic amine which is obtained as an yellow color. The isomeric ratio (E: Z) in the purified product is approximately 90:10 based on 1 H NMR analysis.

Preparation of pyrophosphoramidate of (E) -3-methyl-4- (tetrahydro-2H-pyran-2-yloxy) but-2-enyl Disodium pyrophosphate (0.25 mmol, 1 equivalent) and (E) -3-methyl- are dissolved. 4- (Tetrahydro-2H-pyran-2-yloxy) -but-2-en-l-amine (46 mg, 0.25 mmol, 1 equivalent) in 3 ml of a 1/1 (v / v) mixture of deionized water / acetonitrile and introduced into a glass reaction vessel. 12.5 ml of a 0.2 M solution of l-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride in deionized water / acetonitrile (2.5 mmol, 10 equivalents) are added dropwise using a syringe, while stirring at room temperature . The progress of the reaction is then monitored by ion chromatography (HPAEC). After about 3 hours the reaction mixture is concentrated until a final volume of approximately 5 ml at reduced pressure and the raw material obtained in this stage is converted to the ammonium form by passing the aqueous solution through a column containing an excess of resin DOWEX 50WX8-200 (NH + form) eluting with two column volumes of deionized water. This treatment also allows the unreacted carbodiimide and the cationic by-products to be removed from the reaction medium. The collected solution is then concentrated to a final volume of approximately 5 ml by evaporating the water under reduced pressure (20 mbar - 40 ° C) and used directly for the next step.

Preparation of (E) -4-hydroxy-3-methylbut-2-enyl pyrophosphoramidate (N-HDMAPP) The complete removal of the tetrahydropyranyl protecting group (THP) is achieved by passing the crude solution obtained in the previous step through a column containing (10 g - 24 meq) DOWEX 50 WX8-200 resin (H + form) eluted with two column volumes of deionized water. The resulting acid solution is collected in a 50 ml Falcon® tube placed in an ice bath. The resulting solution is immediately neutralized to pH 8 with 10% ammonium hydroxide solution. The product is then purified by anion exchange chromatography through a 5 g Sep-Pak Accell Plus QMA cartridge (Waters®) eluted in succession respectively with 5 mM, 10 M, 25 mM, 50 mM solutions, 75 mM, 100 mM and 200 mM aqueous bicarbonate, monitoring the fractions eluted by ion chromatography (HPAEC). The fractions corresponding to the purified product are then combined and lyophilized in order to eliminate the ammonium bicarbonate. The isomeric relationship (E: Z) in the purified product is 87:13 based on the HPAEC analysis. Pyrophosphoramidate of (E) -4-hydroxy-3-methylbut-2-enyl is obtained by chromatographic purification (HPAEC) through a column IonPac® AS11, combining multiple chromatographic passes. In order to carry out the biological evaluation, neutral aqueous solutions of the product are sterilized by filtration through a 0.2 μm filter and stored at -20 ° C. If the evaluation is carried out in vivo, the solutions are passed through a column of cationic resin DOWEX 50WX8-200 (Na + form) eluted with two column volumes of deionized water. The synthesis of (E) -4-hydroxy-3-methylbut-2-enyl pyrophosphoramidate (N-HDMAPP) is carried out according to the following reaction scheme. For each step of this synthesis scheme the following references can be used for additional guidance: Step 1: Hecht et al., Tetrahedron Letters, 43 (2002) 8929-8933; Step 2: Miyashita et al, J. Org. Chem., 42 (1977) 3772-3774, Solladié et al, J. Org. Chem. 1993, 58, 2181-2185, and Marshall et al, J. Org. Chem. 1985, 50 (10), 1602-1606; Step 3: Deslongchamps et al, Can. J. Chem. 1979, 57, 3262-3271; Step 4: Coperet et al, Tetrahedron 1996, 52 (35) 11520-11544; Step 5: Sato et al, Chem. Pharm. Bull, 38 (8), 2287-2289 (1990); and Step 6: Miyashita et al, J. Org. Chem., 42 (1977) 3772-3774 (only deprotection reaction). 3 eq. of NaN3 / pentane TBAHS04 / H20 1) Dowex 50WX8 resin (form H +) 2) NaOH or Dowex 50WX8 resin (Na + form) EXAMPLE 2 Production of 3-methylbut-3-enyl pyrophosphorus idato (N-IPP) Preparation of 3-methyl-3-buten-1-yl tosylate Magnesium stirring (4.8 g, 25 mmol) and 4- (N, N-dimethylamino) pyridine (3.4 g, 27.5 mmol) are mixed with magnetic stirring. 90 ml of anhydrous dichloromethane in a three-neck 250 ml flask which is cooled in an ice bath. A solution of 3-methyl-3-buten-1-ol (2.2 g, 25 mmol) in approximately 10 ml of anhydrous dichloromethane is then slowly introduced via a syringe through a septum into the flask, and then the bath is removed. of ice. The reaction is monitored by TLC with silica gel (pentane / ethyl acetate, 85:15 (v / v)). After 2 hours with constant stirring, the mixture is precipitated by dilution in 1 liter of hexane and filtered, and the filtrate is concentrated under reduced pressure. This filtration / suspension step is repeated using diethyl ether, and the resulting oil is purified by flash chromatography on silica gel (pentane / ethyl acetate, 85:15 (v / v)), which produces a yellow oil of 3-methyl-3-buten-l-yl-tosylate (5.6 g, 23.5 mmol, 94% yield) which is kept under dry N2 at 4 ° C.

Preparation of 4-azido-2-methyl-l-ene To a solution of 2.0 g (8.32 mol) of 3-methyl-3-buten-l-yl-tosylate in 20 ml of DMSO is added 820 mg (12.48 mmoles) of sodium azide (NaN3) and 125 mg (catalytic amount) of Nal. The reaction mixture is stirred at 55 ° C overnight. The reaction mixture is cooled to room temperature and 120 ml of water are added. The solution is extracted with 3 x 100 ml of Et20. The combined organic phases are washed with 100 ml of water, 100 ml of brine, dried over Na 2 SO, filtered and concentrated by evaporation. 300 mm Hg at room temperature. 875 mg (7.87 mmol, 95% yield) of 4-azido-2-methyl-1-ene are isolated as a brown oil.

Preparation of 3-methyl-3-en-l-amine A solution of 500 mg (4.50 mol) of 4-azido-2-methyl-1-ene, 3.92 g (14.95 mmol) of the same is stirred at room temperature overnight. Triphenylphosphine PPh3 and 2.7 ml of water in 27 ml of THF and the solvent is evaporated. The resulting crude product is then purified by chromatography with silica gel using CH2Cl2 / MeOH / Et3N 9/1 / 0.5) as eluent. 125 mg (1.47 mol) of 3-methyl-3-en-1-amine are isolated as a yellow oil with 32% isolated yield. The purified amine is kept under dry N2 and stored at -20 ° C for the next step.

Preparation of 3-methylbut-3-ene pyrophosphoramidate (N-IPP) 3-Methylbut-3-enyl pyrophosphoramidate is prepared following the procedure reported in Example 1 for the preparation of 3-methyl-4- (tetrahydroxy) pyrophosphoramidate 2H-pyran-2-yloxy) but-2-enyl: Disodium pyrophosphate (0.25 mmol, 1 equivalent) and 3-methyl-3-en-l-amine (0.25 mmol 1 equivalent) are dissolved in 3 ml of a mixture of 1 / 1 (v / v) deionized water / acetonitrile and introduced into a glass reaction vessel. 12.5 ml of a 0.2 M solution in deionized water / acetonitrile of l-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (2.5 mmoles 10 equivalents) are added dropwise using a syringe, while stirring at room temperature. The pH is adjusted if necessary and maintained in the range of 6-6.5 with the addition of aqueous 0.1 N HCl. The progress of the reaction is then monitored by ion chromatography (HPAEC). After about 3 hours, the reaction mixture is concentrated to a final volume of about 5 ml under reduced pressure and the raw material obtained in this step is converted to the ammonium form by passing the aqueous solution through a column containing an excess of DOWEX 50WX8-200 resin (NH + form) eluted with two column volumes of deionized water. Purification of the crude solution is achieved by anion exchange chromatography through a Sep-Pak Accell Plus QMA cartridge (Waters®) following the procedure reported in Example 1 for the preparation of N-HDMAPP. The syntheses of N-IPP, pyrophosphoramidate of 5-bromo-4-hydroxy-4-methylpentyl (N-BrHPP) (example 3) and N-EpoxPP (example 4) are carried out according to the following reaction scheme. For each step of this synthesis scheme the following references can be used for additional guidance: Step 1: Davisson et al., J. Org. Chem., 1986, 51, pgs. 4768-4779; Step 2: Grieco et al, Tetrahedron 1986, 42 (11), 2847-2853, and Sahasrabudhe, K. et al., J. Am. Chem. Soc. 2003; 125 (26); 7914-7922; Step 3: Brettle R. et al., Bioorg. Med. Chem. Lett. , vol. 6, p. 291 (1996); Step 4: Sato et al, Chem. Pharm. Bull, 38 (8), 2287-2289 (1990); Step 5: Espinosa, et al, (2001a) J Biol Chem 276, 18337-18344; and Step 6: International patent publication no. WO 00/012519. 2 eq. of PPh3 H20 / THF, t.amb. 1) Br2, H20; Initial pH < 7 2) DowexNa resin N-EpoxPP N-BrHPP EXAMPLE 3 Production of 5-bupto-4-hydroxy-4-methylpentyl pyrophosphoramidate (N-BrHPP) As illustrated in the following synthesis scheme, the N-BrHPP compound can be prepared starting from the N-IPP compound described in Example 2 by addition of bromine water to the alkene functional group followed by a neutralization in DOWEX 50WX8-200 resin ( Na + form). The formation of the functional group bro ohydrin with subsequent purification of the crude product can be carried out in accordance with the experimental protocol provided in WO 00/012516 for the preparation of 3- (bromomethyl) -3-butanol-1-yl diphosphate. (BrHPP) or as described in the document by Espinosa et al, J Biol Chem, 276, (2001) 18337-18344. 2 eq. of PPh3 H20 / THF, t.amb. 1) Br2, H20; Initial pH < 7 2) DowexNa resin N-EpoxPP N-BrHPP EXAMPLE 4 Production of 2-2-methyloxyran-2-yl) ethyl pyrophosphoramidate (N-EpoxPP) As illustrated in the following synthesis scheme, the compound N-EpoxPP can be prepared starting from the compound N-BrHPP described in example 3 by treatment with 1M ammonium hydroxide solution (epoxidation reaction) followed by a cation exchange step in resin DOWEX 50WX8-200 (Na + form). The epoxidation reaction with subsequent purification of the crude product can be carried out according to the experimental protocol provided in WO 00/012519 for the preparation of 3,4-epoxy-3-methyl-1-butyl diphosphate (EpoxPP). 2 eq. of PPh3 H20 / THF, t.amb. 1) Br2, H20; Initial pH < 7 2) DowexNa resin N-EpoxPP N-BrHPP EXAMPLE 5 Response to in vitro and in vivo dosing for the compound N-HDMAPP Cytokine release test Cells are thawed (primary polyclonal V? 9Vd2 human cells that have expanded in vitro and stored in the frozen state on days 12-15 of expansion) and are rinsed twice and centrifuged. After removing the supernatant and suspending the cells again, the cells are incubated for 24 hours at 37 ° C in the presence of 100 IU / ml of IL2 (final concentration). The cells are washed and centrifuged, after which the supernatant is removed and the cells are resuspended and adjusted to the appropriate final concentration. The cells are added to the cavities of a 96-well plate. To a row of cavities is added a series of standard dilutions of 3- (bromomethyl) -3-butanol-1-yl diphosphate (BrHPP). The compounds to be evaluated, in this case (E) -4-hydroxy-3-methyl-2-butenyl pyrophosphate (HDMAPP) and the N-HDMAPP compound of the invention are added to the experimental cavities, after several dilutions. The filled plates are incubated 24 hours at 37 ° C to stimulate the cells? D with the test compound and reference compounds, in this case N-HDMAPP, BrHPP and HDMAPP, as described further below. After this time, 100 μl of culture supernatant is taken for dosing of TNFa. The measurement of the dose of TNFa released is carried out as described in the manufacturer's instructions in the kit for immunological test of TNFa enzyme (ref 11121, Immunotech-Beckman Coulter). The OD is read at 405nm, the DO is proportional to the concentration of TNFa released in the culture supernatant. The data is processed with Excel software to compare the concentration of the test compound against the concentration of TNFa and for the calculation of the EC50 for each test compound.

Bioactivity of N-HDMAPP in vitro The bioactivity of the N-HDMAPP compound is evaluated using a TNFα release test as described above. Figure 1 shows the in vitro activity. The BrHPP and HDMAPP compounds are included for comparison purposes. The EC50 in vitro is then evaluated in this in vitro relative selection test, in which the above tests with cells calibrated using a standard composition of BrHPP present an EC50 of approximately 15 nM for BrHPP. As will be appreciated, any other appropriate tests such as cell amplification can be used in the evaluation of the compounds. It is determined that the EC50 for N-HDMAPP is 0.63 nM while the EC50 in vitro for HDMAPP is 2.1 nM and the EC50 in vitro for BrHPP is 37.7 nM. Because the test provides a relative result instead of an absolute EC50 value, the results indicate that the N-HDMAPP compound has 3-4 times more potency than the more potent compounds analyzed so far. The maximum level of TNFa release is also evaluated. As shown in Figure 1, the compound N-HDMAPP produces a greater maximum TNFa release than with the other compounds analyzed. Although the other compounds evaluated differ in potency (EC50), these show similar levels of maximum TNFa release - between 1200 and 1500 pg / ml approximately of TNFa release. However, the N-HDMAPP compound produces a statistically significant increase in the maximum TNFα release of more than about 1800 pg / ml of TNFa release, suggesting that N-HDMAPP may lead to increased TV cell activation? 9Vd2 absolute in vivo than that which can be obtained at any concentration of the other compounds.

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Claims (22)

NOVELTY OF THE INVENTION Having described the present invention, it is considered as a novelty and therefore the content of the following is claimed as property: CLAIMS

1. - A T? D cell activator of the formula (I): Formula (I) characterized in that Cat + represents a cation or several cations, identical or different, organic or mineral (including proton); m is an integer from 1 to 3; B is O, NH, or any group that can be hydrolyzed; Y = O'Cat +, an alkyl group of C? -C3, a group -AR, or a radical that is selected from the group consisting of a nucleoside, an oligonucleotide, a nucleic acid, an amino acid, a peptide, a protein, a monosaccharide, an oligosaccharide, a polysaccharide, a fatty acid, a lipid simple, a complex lipid, a folic acid, a tetrahydrofolic acid, a phosphoric acid, an inositol, a vitamin, a co-enzyme, a flavonoid, an aldehyde, an epoxide and a halohydrin; A is 0, NH, CHF, CF2 or CH2; and R is a linear, branched, or cyclic, aromatic or non-aromatic, saturated or unsaturated C? -C50 hydrocarbon group, optionally interrupted by at least one heteroatom, wherein said hydrocarbon group comprises an alkyl, an alkylenyl, or an alkynyl, preferably an alkyl or an alkylene, which may be substituted with one or more substituents which are selected from the group consisting of: an alkyl, an alkylenyl, an alkynyl, an epoxyalkyl, an aryl, a heterocycle , an alkoxy, an acyl, an alcohol, a carboxylic group (-C00H), an ester, an amine, an amino group (-NH2), an amide (-C0NH2), an imine, a nitrile, a hydroxyl (-0H), an aldehyde group (-CH0), a halogen, a halogenoalkyl, a thiol (-SH), a thioalkyl, a sulfone, a sulfoxide, and a combination of them.

2. - The T? D cell activator according to claim 1, characterized in that said activator is a compound of the formula (X): (X) wherein R3, R, and R5, identical or different, are a hydrogen or an alkyl group of C? ~ C3, W is -CH- or -N-, R6 is a C2-C3 acyl, an aldehyde, an alcohol of C? ~ C3, or a C2-C3 ester, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), B is 0 or NH, m is an integer of 1 to 3, and Y is 0"Cat +, a nucleoside, or an -AR radical, in which A is 0, NH, CHF, CF2 or CH2, and R is selected from the group consisting of 1), 2), or 3)

3. The T? d cell activator according to claim 2, characterized in that said activator is a compound of the formula (XI).

4. - The T? D cell activator according to claim 3, characterized in that said activator is a compound of the formula (XII) (XII) N-HDMAPP

5. - T cell activator according to claim 1, characterized in that said activator is a compound of formula (II): wherein X is a halogen (which is preferably selected from I, Br and Cl), B is 0 or NH, m is an integer from 1 to 3, Rl is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different, organic or mineral (including the proton), and n is an integer from 2 to 20, and Y is 0"Cat +, a nucleoside, or an -AR radical, in which A is 0 , NH, CHF, CF2 or CH2 and R is selected from the group consisting of 1), 2), or 3)

6. The T cell activator according to claim 5, characterized in that said activator is a compound of the formula (III)

7. - The T? D cell activator in accordance with claim 5, characterized in that said activator is a compound of the formula (V): OH O or Br- -C- - (CHo) o- -N- I PI-0- -p-O, xCat H, H CH, 0 ~ O (V) N-BrHPP

8. - The T? D cell activator in accordance with claim 1, characterized in that said activator is a compound of the formula (VI): in which R1 is a methyl or ethyl group, Cat + represents a cation or several cations, identical or different organic or mineral (including the proton), B is 0 or NH, m is an integer from 1 to 3, and n is an integer from 2 to 20, and Y is 0"Cat +, a nucleoside, or an -AR radical, in which A is 0, NH, CHF, CF2 or CH2 and R is selected from the group consisting of 1), 2), or 3). - A pharmaceutical composition comprising a T? D cell activator according to any of claims 1-8.10- The use of a T? D cell activator according to any of claims 1-8 for the manufacture of a pharmaceutical composition for regulating T? d cells in a human subject 11. The use of a T? d cell activator according to any of claims 1-8 for the manufacture of a pharmaceutical composition for treating an individual. who suffers from or who is susceptible to suffering from cancer, an infectious disease, an to-immune or an allergic disease. 12. The use according to claim 11, characterized in that said cancer is a solid tumor. 13. The use of a T cell activator according to any of claims 1-8 as a adjuvant for vaccine. 14. A vaccine composition comprising a T? D cell activator according to any of claims 1-8 as a vaccine adjuvant. 15. A method for preparing a diphosphoramidate monoester compound comprising: (a) reacting an alkyl halide R-X in a coupling step with a reagent of diethylphosphoramidate or diethylchlorophosphate type; (b) reacting the compound prepared in step (a) in a saponification step with which the O-ethyl groups are removed; and (c) reacting the compound prepared in step (b) in a phosphorylation step with which a diphosphoramidate monoester is prepared, characterized in that R is a linear, branched, or cyclic C1-C50 hydrocarbon group, aromatic or non-aromatic, saturated or unsaturated, optionally interrupted by at least one heteroatom, wherein said hydrocarbon group comprises an alkyl, an alkylenyl, or an alkynyl, preferably an alkyl or an alkylene, which may be substituted with one or various substituents which are selected from the group consisting of: an alkyl, an alkylenyl, an alkynyl, an epoxyalkyl, an aryl, a heterocycle, an alkoxy, an acyl, an alcohol, a carboxylic group (-COOH), a ester, an amine, an amino group (-NH2), an amide (-CONH2), an imine, a nitrile, a hydroxyl (-0H), an aldehyde group (-CHO), a halogen, a halogenoalkyl, a thiol ( -SH), a thioalkyl, a sulfone, a sulfoxide, and a combination thereof, and characterized in that X is a portion that can be displaced by a diethylphosphoramidate group under appropriate conditions. 16. The method according to claim 15, characterized in that X is a group NH2 and said compound R-X is reacted in a coupling step with a diethylchlorophosphate compound. 17. The method according to claim 15, characterized in that X is selected from the group consisting of I, Br and Cl. 18. A method for preparing a compound (E) -2- (4-azido-2-methylbut-2-enyloxy) tetrahydro-2-t-pyran, which comprises providing a compound (E) -2- (4-chloro- 2-methylbut-2-enyloxy) tetrahydro-2i? -pyran and reacting said compound with a sodium azide in a biphasic mixture of water-pentane in the presence of phase transfer catalyst. 1

9. A method for activating a T? D cell, the method comprises placing a T? D cell in contact with a T? D cell activator according to any of claims 1-8. 20. The method according to claim 19, characterized in that the T? D cell is contacted with said cell activator T? D in vi tro. 21. A T? D cell activated according to a method according to claim 19 or 20. 22. The use of a T? D cell according to claim 21 for the manufacture of a pharmaceutical composition.