EP0756603A1 - Multirepresentation d'un analogue peptidique du substrat de la dppiv, notamment de type kpr, afin d'inhiber l'entree du hiv dans les cellules - Google Patents

Multirepresentation d'un analogue peptidique du substrat de la dppiv, notamment de type kpr, afin d'inhiber l'entree du hiv dans les cellules

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Publication number
EP0756603A1
EP0756603A1 EP95918043A EP95918043A EP0756603A1 EP 0756603 A1 EP0756603 A1 EP 0756603A1 EP 95918043 A EP95918043 A EP 95918043A EP 95918043 A EP95918043 A EP 95918043A EP 0756603 A1 EP0756603 A1 EP 0756603A1
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EP
European Patent Office
Prior art keywords
hiv
molecule according
kpr
tass
inhibition
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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Application number
EP95918043A
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German (de)
English (en)
French (fr)
Inventor
Ara Hovanessian
Christian Callebaut
Bernard Krust
Etienne Jacotot
Sylviane Muller
Jean-Paul Briand
Gilles Guichard
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Institut Pasteur de Lille
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Institut Pasteur de Lille
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Publication of EP0756603A1 publication Critical patent/EP0756603A1/fr
Withdrawn legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2740/00—Reverse transcribing RNA viruses
    • C12N2740/00011—Details
    • C12N2740/10011—Retroviridae
    • C12N2740/16011—Human Immunodeficiency Virus, HIV
    • C12N2740/16111—Human Immunodeficiency Virus, HIV concerning HIV env
    • C12N2740/16122—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes

Definitions

  • the present application relates to inhibitors of infection by an HIV retrovirus, comprising a peptide analog of the DPPIV substrate or / and of motifs conserved in the V3 loop of the envelope glycoprotein of a HIV type retrovirus.
  • HIV human immunodeficiency virus
  • AIDS acquired immunodeficiency syndrome
  • retroviruses So far two distinct but related types of retroviruses, the HIV-1 type and the HIV-2 type have been identified.
  • Infection of CD4 + T lymphocytes, onocytes and macrophages by HIV requires the interaction of the virus env gene products with the cellular CD4 receptor (Klatzman D, et al. Nature 1984; 312: 767-768).
  • the HIV env gene codes for a precursor polyprotein which is cleaved by a cellular protease, leading to the production of extracellular or surface (SU) and transmembrane (TM) glycoproteins (Eiden LE, et al. Immunol. Today 1992 13 : 201-206).
  • the surface protein through non-covalent interactions with the transmembrane protein, is exposed on the surface of infected cells or on the surface of viral particles.
  • the CD4 receptor is a glycoprotein composed of an amino-terminal extracellular region containing four domains related to the immunoglobulin family and of a carboxy-terminal region containing the transmembrane and cytoplastic domains (Eiden et al).
  • the first a-terminal domain of the CD4 molecule appears to be the main binding site for HIV surface proteins. Consequently, this bond induces a change in conformation of the SU-TM complex (complex formed by surface and transmembrane glycoproteins) allowing the interaction of the amino-terminal domain of the transmembrane protein with the cell membrane, thus causing the fusion of viral and cell membranes (Marsh M, et al Immunol.
  • V3 loop contains 35 to 41 amino acids whose cysteines at the two ends are linked, by a disulfide bridge forming a loop.
  • the V3 loop contains conserved basic residues and in addition the sequence near the bound cysteines is similar in most isolates.
  • the inventors are interested in a particular sequence of the V3 loop of the surface glycoprotein of the envelope of the HIV-1 retrovirus, distinct from those which have been the subject of experiments in the prior art, and in the corresponding sequences.
  • HIV-2 and SIV retroviruses They observed that a very large number of identified isolates of HIV, in particular HIV-1 (more than 90%) exhibit a peptide motif constituted by the amino acid sequence Gly-Pro-Gly (Glycine - Proline - Glycine or according to the one-letter code GPG) and more particularly a Gly-Pro motif, preserved. This GP or GPG motif also remains unchanged in vivo for several years in individuals infected with HIV-1, while other substitutions appear in the V3 loop (Holmback et al. J. Virol. 1993, 67: 1612-1619).
  • the inventors have demonstrated that a surface cellular protein interacts with the glycoproteins of the HIV or SIV envelope, in particular at the level of the V3 loop and is capable of cleaving it. This cleavage would take place at a site not identified as such until the present invention.
  • the surface cellular protein is dipeptidyl peptidase IV (DPPIV) also called antigen or CD26 receptor (Barclay AN, et al. (1993). Facts book. London: Harcourt Brace Jovanich).
  • DPPIV / CD26 is also called: DPIV, DAPIV, SGP-115/107, Tal and Tal03.
  • DPPIV is a cell surface protease (EC 3.4.14.5) which generally has exopeptidase activity and under certain conditions endopeptidase activity (Nagatsu T, et al. Anal. Bioche. 1976; 74: 466-476 - Kudo M, et al J. Bioche. 1985; 97: 1211-1218 - Kenny AJ, et al. Biochem. J. 1976; 155: 169-182).
  • X can be an amino acid residue, in particular Glycine (Gly), Alanine (Ala), Lysine (Lys), Arginine (Arg), Glutamic Acid (Glu), Aspartic Acid (Asp) or even Proline (Pro).
  • the CD26 antigen is widely expressed as a 110 to 120 kDa transmembrane glycoprotein, which is identical to the cell surface serine protease called dipeptidyl peptidase IV.
  • This enzyme which is generally considered as an exopeptidase, cleaves dipeptides in the amino-terminal part of polypeptide substrates, since the penultimate residue is a proline residue; that is to say, this enzyme allows the cleavage of X-P-polypeptide, P being a Proline residue, when X is a free N-terminal amino acid (Kudo M, et al. J. Biochem. 1985; 97 : 1211-1218).
  • DPPIV has been described as capable of catalyzing an "endopeptidase-like" cleavage, after a "GP-like” dipeptide motif within synthetic peptides (Kenny AJ, et al. Biochem. J. 1976; 155: 169 -182) and to bind to collagen (Beauvois B, et al. I. Marcel Dekker (Eds.) New York, 1991). DPPIV binds collagen but this binding of DPPIV to collagen does not affect its enzymatic activity (Hanski C, et al. Exp. Cell Res. 1988; 178: 64-72), thus suggesting the existence of attachment and catalytic in different areas of DPPIV / CD26.
  • DPPIV is a glycoprotein comprising several oligosaccharide chains linked to N residues.
  • This enzyme exists in a wide variety of tissues including in lymphoid and non-lymphoid organs (Gorrel MD, et al. Cell.
  • DPPIV is a co-receptor for HIV, which is capable of binding the V3 loop, and / or other regions in the glycoproteins of the SU and TM viral envelope. This step concerning the interaction of the SU / TM complex with DPPIV / CD26 is essential for the penetration of HIV into CD4 T cells.
  • the subject of the invention is therefore inhibitors of infection by an HIV retrovirus, comprising a plurality of motifs capable of being recognized by an ectoprotein (on the surface of cells) in particular by the CD26 receptor (also called the DPPIV enzyme). reasons all being carried by a peptide matrix or not authorizing their multiple presentation to the enzyme. In what follows, reference will be made to all of these motifs thus presented to the enzyme under the expression "repeated motifs”.
  • the matrix allowing the multiple presentation of a chosen motif is defined in relation to its ability to make the above motif accessible.
  • peptide (s) as used in the present context, whether it is “repeating peptide motifs" or the “peptide matrix”, is understood to cover both purely peptide molecules (in so far as they are based on natural amino acid residues) and molecules derived from the preceding ones by replacing all or part of its natural amino acid residues by optical isomers of these amino acids natural or even by "non-peptide” groups (leading for example to "pseudo-peptides”), provided that these replacements do not entail a significant alteration of the relevant properties of said repeating units (in particular their ability to be recognized by the DPPIV) or matrix (in particular capacity for presentation of said repeating motifs to the enzyme).
  • the peptide matrix carrying the "repeated motifs" contains amino acid residues having a reactive function which is not engaged in a peptide bond with neighboring amino acid residues, these reactive functions involved in fixing said repeated patterns on this matrix.
  • the peptide matrix comprises amino acid residues of lysine type (K), the aforementioned repeat units then being grafted onto the epsilon-a groups ino of at least part of these lysine residues.
  • amino acid residues can also be used for the grafting of the aforementioned repeating units, such as arginine or also glutamyl residues which carry a carboxyl function not engaged in peptide bonds with other amino acids of the matrix.
  • the matrix can be synthetic and therefore prepared by chemical synthesis, in particular by using solid phase synthesis as described by Merrifield.
  • a first molecule of interest in the context of carrying out the invention is, for example, characterized in that the peptide matrix comprises the following amino acid sequence: K X., KX 2 X 3 in which X : is preferably , lysine, valine, alanine or glutamic acid or isoleucine, X 2 and X 3 represent glycine or proline and are different from each other.
  • the repeating units can in this case be linked to the e-amino functions of the lysine residues, in particular by peptide bonds, and can be presented in the same plane or on the contrary in different planes. Likewise, these reasons can be linked same side of the matrix with respect to the lysine residue or in opposite manner.
  • amino acid residue X ⁇ can, if necessary, be deleted.
  • peptide matrices of interest for the production of the molecules of the invention include an amino acid chain chosen from the following sequences
  • the amino acids contained in the peptide matrix are in the dextrorotatory form.
  • the peptide bonds between the amino acids of the matrix are modified in particular to improve their resistance to hydrolysis by proteases in vivo.
  • the number of repeat units linked to the lysine residue of the matrix can vary considerably.
  • This number can be determined according to the number of lysine residues present on the peptide matrix and can be adapted so as to optimize their capacity to inhibit the entry of the virus into lymphocytes in vivo, and taking into account the influence of presence of an increased number of patterns on the possible in vivo toxicity of the molecules thus obtained.
  • the number of repeated patterns presented by the matrix is between 2 and 8, preferably between 4 and 6 patterns, for example 5 patterns.
  • the number of amino acids contained in the repeated motif will be taken into account to determine the number of motifs linked to the peptide matrix.
  • modifications of the bonds between the amino acids of the repeating units and / or of the peptide matrix can be of different natures.
  • a suitable modification can be carried out at the level of the peptide bond, in order to prevent the cleavage of this bond and therefore in order to decrease or suppress the sensitivity to peptidases.
  • D enantiomers may be preferred in the context of the invention rather than molecules having an L configuration.
  • these two charges will be separated by at least one neutral amino acid residue such as Gly, Pro, Ala.
  • the patterns will for example have as sequence, RP, KP, PK, PR, RK, KR, KPR, RPK, PKR, PRK, KER, KGQ, KGR, RPR, RPG, AHxPR, pyrKR, RPGR, KPGR, KPRG, GPGR, IPIG, GPGRAF, KRPGNK, RPGNK, KRPRQ, KPRQAG
  • a particularly preferred molecule (involving only "real" peptide bonds) for carrying out the invention corresponds to the following formula: RRRR
  • the repeated patterns linked to the lysine residues of the peptide matrix can be similar or of different nature.
  • repeating units can be chosen such that they comprise both a B epitope and a T epitope of the external envelope glycoprotein of an HIV retrovirus.
  • the molecule is characterized in that the peptide matrix is replaced by a functionally equivalent molecule chosen from polyamines, for example Tris (2-aminoethyl) amino or dendrimers (R. olf , Frequency Chemistry p. 13-17, Large molecules).
  • a functionally equivalent molecule chosen from polyamines, for example Tris (2-aminoethyl) amino or dendrimers (R. olf , Frequency Chemistry p. 13-17, Large molecules).
  • the subject of the invention is moreover a composition capable of inhibiting infection due to a human retrovirus of HIV type, in particular of HIV-1 or HIV-2 type, characterized in that it comprises, as active principle, a molecule above defined.
  • Such a composition is in particular defined by its capacity to inhibit the interaction between the CD26 cell receptor (DPPIV) and the external envelope glycoprotein of a HIV-type retrovirus, this interaction manifesting the inability then acquired by the retrovirus to penetrate into CD4 lymphocytes, as evidenced by the absence of syncitia formation and induction of apoptosis, as observed in control cultures.
  • DPPIV CD26 cell receptor
  • the mechanism of action of the inhibitors according to the invention does not normally involve an inhibition of the enzymatic activity of DDPIV, which is normally not affected.
  • the invention also relates to the use of a molecule as defined above for the preparation of a medicament for the treatment of an infection due to a human retrovirus of HIV type, in particular by preventing the fusion of cell membranes.
  • composition characterized in that it comprises, as active principle, a molecule as defined above.
  • This pharmaceutical composition can be used for the treatment or prevention of infection with the HIV retrovirus, in particular as a munogenic composition, in particular as a vaccine.
  • TASP type template assembled synthetic protein
  • the matrix is used to form a covalent anchoring region of a peptide (for example the peptide KPR) which inhibits, in time that monomer at a relatively high molarity (10 M), the entry of HIV into CD4 + cells.
  • a peptide for example the peptide KPR
  • DCM Dichloromethane
  • TFA trifluoroacetic acid
  • N, N-dimethylformamide (DMF) and diisopropylethylamine (DIEA) were purchased from SDS (Peypin, France).
  • Tert-butyl-methyl-ether MTBE
  • benzotriasolyl-oxy-tris dimethylamino
  • BOP reagent benzotriasolyl-oxy-tris
  • HOBt hydroxybenzotriazole
  • the protected amino acids come from Neosystem (Strasbourg, France).
  • the PAM Boc Cys resin (4 Me Bzl) comes from A.B.I. (Roissy, France).
  • Hydrogen fluoride comes from Prodair (Strasbourg, France).
  • the protective side chain groups used were: the cyclohexyl ester (Glu 4), the chlorobenzyloxycarbonyl (Lys 9), the t-butyloxycarbonyl (Lys 3, 5, 8).
  • the lysine at position 10 was introduced in the form of a Boc Lys Boc derivative.
  • the cycle used for the incorporation of each amino acid derivative is as follows:
  • the Boc protecting groups of the matrix PAM resin were cleaved in 60% TFA / DCM for 1 min (prewash) and 30 min, then the resin was washed with DCM and DMF.
  • the new resin substitution was 3 meq / g (0.5 mmol of amino groups in the reaction vessel).
  • the elongation of the peptide chains was obtained using 4 eq in excess of Fmoc Arg (Pmc), Fmoc Gly and Fmoc Lys (Boc).
  • the deprotection time of the Fmoc protection groups was increased to 3 ⁇ 7 min in 25% of a piperidine / DMF mixture.
  • the resin was washed with DCM and dried under nitrogen.
  • the resin-bound peptide (620 mg) finally obtained was tested with 10 ml of HF and 1 ml of anisol at 0 ° C for 45 min. After removal of the hydrogen fluoride in vacuo, the resin was washed with ether and the peptide was eluted with 8 ml of pure TFA and precipitated again with MTBE. After centrifugation, the centrifugation pellet was dissolved in 10 ml of water and lyophylized. f) Analysis and purification of the molecule The homogeneity of the final product was checked by means of an analytical step on a reverse phase column of the C18 nucleoside type (150 x 4.6 mm) using a TEAP buffer system. The linear gradient detected at 201 nm was 1 to 21% in 20 min.
  • ALBA 2 Lys (Z (N0 2 ) -pyrrolidid HCL K: Lys; P: Pro; R: Arg Compared to ALBA 2, molecule 5 (KPR) TASS is 5,000 to 10,000 times less active for the inhibition of DPP IV activity.
  • HIV-1 LAI The entry of HIV-1 LAI was tested by infection of CEM cells for 1 h. The extracellular virus was eliminated by treatment with trypsin and the internalization of HIV was followed by testing the concentration of p25 (Callebaut et al, 1993, cited above).
  • molecule 5 (KPR) TASS is a powerful inhibitor of the entry of HIV.
  • the IC 50 value is approximately 10 ⁇ M.
  • 5 (KPR) TASS is 500 times more active. It should be noted that at 200 ⁇ M of 5 (KPR) TASS, the DPP IV enzymatic activity is not affected at all.
  • CEM cells were incubated (37 ° C, 30 min) with different concentrations (200, 100, 50, 25 and 5 ⁇ M) of 5 (KPR) TASS before infection with HIV-1 LAI.
  • the cells infected in the presence of the inhibitor were then cultured for 4 days and the production of virus was tested by monitoring the concentration of p25 in the culture supernatant.
  • the production of HIV was considerably inhibited by the molecule 5 (KPR) TASS and this inhibition was dependent on the dose: more than 90% inhibition at 20 ⁇ M and around 50% inhibition at 5 to 10 ⁇ M .
  • 5 (KPR) TASS As the production of the virus was inhibited by 5 (KPR) TASS, the formation of syncytia was also inhibited.
  • 5 (KPR) TASS is an entry inhibitor, as demonstrated in section c (i.e. by inhibiting fusion of viral and cell membranes) and further show that 5 (KPR) TASS is also an inhibitor of syncytia formation by preventing fusion of the membranes of infected cells with neighboring cells.
  • the molecule 5 (KPR) TASS has no toxic effect on cell growth
  • the cells (CEM clone 13; CEM 174, HUT 78) were incubated (37 "C, 30 min) with different concentrations (80, 40, 20 and 10 ⁇ M) of 5 (KPR) TASS before infection with HIV-2 ROD, HIV-2 EHO and SIV mac.
  • the infected cells in the presence of the inhibitor were then cultured for 4 days, and virus production was tested by observing • 1 reverse transcriptase activity in the culture supernatant.
  • the HIV-2 and SIV production was significantly inhibited by 5 (KPR) TASS, and 50% inhibition doses were estimated to be approximately 10-20 ⁇ M from 5 (KPR) TASS.
  • g) 5 (KPR) TASS inhibits induction of cell death or apoptosis
  • DPP IV activity inhibitors are not necessarily inhibitors of HIV entry
  • ALBA 2 H-Lys- [Z (NO 2 )] -pyrrolidide
  • DPP IV activity at the cell surface is 90% inhibited at 20 ⁇ M and 50% at 1 ⁇ M.
  • ALBA 2 exerts an effect neither on the entry of HIV, nor on the. syncytia formation, nor on the induction of apoptosis, even at a concentration of 50 ⁇ M.
  • the concentration of 100 ⁇ M is toxic to cells.
  • the TASS molecule 5 (KPR) which exerts only a weak effect on the DPP IV activity, considerably blocks the infection of cells by HIV.
  • IC 50 dose of the inhibitor required for 50% inhibition
  • TASS CEM cells were incubated (37 ° C, 30 min) before 1 • infection with HIV-1 LAI with different concentrations (from 50 to 0.5 ⁇ M). The cells infected in the presence of the inhibitor were then cultured for 4 days and the production of virus was evaluated by measuring the concentration of p25 in the culture supernatant.
  • the table below indicates the concentration of the peptide which gives at least 90% and 50% inhibition of the production of the virus.
  • MOLT cells a line of human T lymphocytes (CD4 + and CD26 +) were incubated (5 min at room temperature) with or without the addition of 50 ⁇ M of 5 (KPR) TASS or 50 ⁇ M of ALBA 2 (which inhibits the enzymatic activity but not the entry of the virus).
  • Antibodies were then added such as MAb 1F7 anti CD26, MAb BA5 another anti CD26 antibody (Immunotech) which does not affect the entry of HIV, MAb OKT4 anti-CD4 (Ortho Diagnostics Systems), MAb ALBl anti CD45 ( Immunotech) (tyrosine kinase associated with CD26) and the cells are incubated at 4 ° C for 1 h.
  • FACS Fluorescence-activated cell sorting analysis
  • MAb 1F7 and MAb BA5 see the figure providing the characteristic spectra resulting from the analysis by FACS for the recognition of CD26 (by MAb 1F7 and MAb BA5), CD4 (by MAb OKT4) and CD45 (by MAb ALB1) on the surface of MOLT cells in the absence or presence of 5 (KPR) TASS (cited as S13) and ALBA2.
  • glycoprotein complex forming the envelope of the virus and comprising the surface protein SU (gpl20) and the transmembrane protein TM (gp41), plays an important role in HIV infection; gpl20 contains the CD4 binding site, and gp41 the peptide motif necessary for the membrane fusion process.
  • gpl20 and gp41 forms a conformational complex which is essential for the two main functions of the gpl20 / 41 complex; the entry of the core of the virus into the cells by fusion of the viral and cell membranes, and the induction of the cytopathogenic effect by fusion of the membranes of the infected cells (expressing the gpl20 / gp41 complex on the surface) with those neighboring cells.
  • gpl20 / gp41 complex on the surface of infected cells leads to an interaction with the CD4 molecules of neighboring uninfected cells, which results in a cytopathogenic effect characteristic of HIV (Laurent-Crawford et al., 1993, AIDS Res. Hum Retroviruses _, 761-773): formation of syncytia (multinucleated cells generated by the fusion of infected cells with uninfected cells) and / or cell death by apoptosis (chromatin fragmentation at the level of internucleosomal links).
  • 5 (KPR) TASS is a potential inhibitor of HIV entry and infection. It presents very little effect, even if it can even be observed, on the enzymatic activity DPP IV. Thus it can be thought that 5 (KPR) TASS acts mainly on the recognition site of the substrate (that is to say the V3 loop of gpl20) without affecting the catalytic site of the CD26 molecule.
  • 5 (KPR) TASS acts mainly on the recognition site of the substrate (that is to say the V3 loop of gpl20) without affecting the catalytic site of the CD26 molecule.
  • One of the explanations for the higher activity of 5 (KPR) TASS compared to the monomer KPR could be found in the fact that the molecule CD26 would exist in the form of a multimer hence hence also a more efficient presentation of a multimer more efficient KPR type.
  • the KPR residue in the TASS molecule can also be replaced by other peptides to form variants of this inhibitor, for example: RPK, RPR, PKR, KGR.
  • the tripeptide KPR is one of the motifs conserved in the V3 loop, precisely found in the C-terminal part of the V3 loop of certain HIV-2. Due to the presence of the dipeptide KP, KPR could serve as a substrate for the enzyme DPPIV / CD26. Furthermore, we have shown that the tripeptide KPR monomer at 10 mM inhibits the entry of HIV and also inhibits the activity of DPPIV. On these data, we have shown here that the multimeric presentation of this tripeptide KPR (for example 5 (KPR-TASS) considerably increases the effectiveness of this molecule to inhibit the entry of HIV, the formation of syncytia and the induction of apoptosis.
  • KPR type of peptide 5
  • TASS appear to interact with CD26 in an area that constitutes one epitope of the antibody onoclonal anti-CD26 MAb 1F7.
  • Example of the inhibitory effect of various multimeric synthetic peptides (corresponding to the TASS molecule or to modified forms of the TASS molecule), on HIV-1 LAI infection in CEM cells.
  • CEM cells were incubated (15-30 minutes, 37 ° C) with the different peptides before infection with the HIV-1 LAI strain (60 minutes, 37 ⁇ C). The cells were then centrifuged and suspended in a fresh culture medium containing the various inhibitors. Syncytia formation was followed on days 3 and 4 after infection (post-infection (pi)) and the culture supernatants were tested to determine their concentration of major core protein of HIV, p24 / p25 . A very good correlation has been observed between the inhibition of syncytia formation and the production of HIV (ie the concentration of p24 / p25).
  • the TASS matrix is composed of the following residues: KKKGPKEKGC.
  • KKKGPKEKGC a modified TASS matrix was used in which the proline residue (P) had been modified by a residue G in accordance with the following sequence: KKKGGKEKGC. This last sequence is called TASS (GG).
  • Another modified TASS molecule corresponding to the sequence KKKKGC was also used. In some cases, the MAP matrix was used for comparison.
  • the different peptide-TASS molecules are active at concentrations in micrograms to block the entry of HIV, infection and formation syncytium.
  • the value of the inhibitory concentration at 50% (IC 50 ) for some of the molecules is between 0.5 and 2 ⁇ M.
  • inhibitor motif would contain the following residues:
  • Two amino acids should be basic residues, for example K or R; K can be replaced by R and vice versa. These two basic residues could occupy any one of positions l, 2 or 3 (see example N ⁇ l, 16, 21, 25 and 33).
  • residue P could be deleted or replaced by a residue G to obtain an active motif (N ° 15 and 30) but replacement with a type E residue would reduce the inhibitory action (see N ° 16).
  • the ⁇ -amino group of the motif does not play a crucial role since it can be acetylated (see No. 35) without the inhibitory activity being reduced considerably. Likewise the introduction of an acid pyroglutamigue at position 1 does not affect the inhibitory activity (see No. 27).

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EP95918043A 1994-04-22 1995-04-21 Multirepresentation d'un analogue peptidique du substrat de la dppiv, notamment de type kpr, afin d'inhiber l'entree du hiv dans les cellules Withdrawn EP0756603A1 (fr)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
FR9404895 1994-04-22
FR9404895A FR2719049B1 (fr) 1994-04-22 1994-04-22 Multireprésentation d'un analogue peptidique du substrat de la DPPIV, notamment de type KPR, afin d'inhiber l'entrée du HIV dans les cellules.
PCT/FR1995/000528 WO1995029190A1 (fr) 1994-04-22 1995-04-21 Multirepresentation d'un analogue peptidique du substrat de la dppiv, notamment de type kpr, afin d'inhiber l'entree du hiv dans les cellules

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EP (1) EP0756603A1 (ja)
JP (1) JPH10502337A (ja)
AU (1) AU2412595A (ja)
CA (1) CA2188470A1 (ja)
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WO (1) WO1995029190A1 (ja)

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* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6825169B1 (en) 1991-10-22 2004-11-30 Trustees Of Tufts College Inhibitors of dipeptidyl-aminopeptidase type IV
US5965532A (en) * 1996-06-28 1999-10-12 Trustees Of Tufts College Multivalent compounds for crosslinking receptors and uses thereof
US6011155A (en) * 1996-11-07 2000-01-04 Novartis Ag N-(substituted glycyl)-2-cyanopyrrolidines, pharmaceutical compositions containing them and their use in inhibiting dipeptidyl peptidase-IV
AU7033798A (en) * 1997-03-12 1998-09-29 Centre National De La Recherche Scientifique A novel cell surface receptor for hiv retroviruses, therapeutic and dia gnostic uses
US6040145A (en) 1997-05-07 2000-03-21 Tufts University Potentiation of the immune response
US6100234A (en) * 1997-05-07 2000-08-08 Tufts University Treatment of HIV
ES2285785T3 (es) 1997-09-29 2007-11-16 Point Therapeutics, Inc. Estimulacion de celulas hematopoyeticas in vitro.
AU770319C (en) 1998-05-04 2004-11-25 Point Therapeutics, Inc. Hematopoietic stimulation
AU765370B2 (en) 1998-06-05 2003-09-18 Point Therapeutics, Inc. Cyclic boroproline compounds
CO5150173A1 (es) * 1998-12-10 2002-04-29 Novartis Ag Compuestos n-(glicilo sustituido)-2-cianopirrolidinas inhibidores de peptidasa de dipeptidilo-iv (dpp-iv) los cuales son efectivos en el tratamiento de condiciones mediadas por la inhibicion de dpp-iv
US6890904B1 (en) 1999-05-25 2005-05-10 Point Therapeutics, Inc. Anti-tumor agents
US6172081B1 (en) 1999-06-24 2001-01-09 Novartis Ag Tetrahydroisoquinoline 3-carboxamide derivatives
US6110949A (en) * 1999-06-24 2000-08-29 Novartis Ag N-(substituted glycyl)-4-cyanothiazolidines, pharmaceutical compositions containing them and their use in inhibiting dipeptidyl peptidase-IV
US6617340B1 (en) 1999-07-29 2003-09-09 Novartis Ag N-(substituted glycyl)-pyrrolidines, pharmaceutical compositions containing them and their use in inhibiting dipeptidyl peptidase-IV
ES2339813T3 (es) 2001-06-27 2010-05-25 Glaxosmithkline Llc Fluoropirrolidinas como inhibidores de dipeptidil peptidasas.
WO2006115230A1 (ja) * 2005-04-22 2006-11-02 Credia Japan Co., Ltd. アミノ酸残基又はペプチド残基を有する化合物、及びその製造方法
KR20080033176A (ko) * 2005-05-23 2008-04-16 가부시키가이샤 쿠레디아쟈판 광증감 작용을 갖는 화합물, 광전극 및 광증감형 태양 전지
WO2007061077A1 (ja) * 2005-11-25 2007-05-31 Credia Japan Co., Ltd. 核酸導入用担体、核酸導入用キット、及び核酸の細胞内への導入方法
FR2900341B1 (fr) * 2006-04-27 2012-09-14 Centre Nat Rech Scient Utilisation de ligands synthetiques multivalents de la nucleoline de surface pour le traitement du cancer
WO2009141687A1 (en) 2008-05-22 2009-11-26 Centre National De La Recherche Scientifique (Cnrs) New optically pure compounds for improved therapeutic efficiency
BR112013008026A2 (pt) 2010-10-04 2016-06-21 Centre Nat Rech Scient composições que compreendem ligantes sintéticos multivalentes de nucleolina de superfície e glicosaminaglicanos

Family Cites Families (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
FR2707170A1 (fr) * 1993-06-04 1995-01-13 Pasteur Institut Expression des récepteurs CD4 et CD26 dans des cellules recombinantes, inhibiteurs du récepteur CD26.
IL110929A0 (en) * 1993-09-13 1994-11-28 Armel Sa Multiple branch peptide constructions and pharmaceutical compositions containing them

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See references of WO9529190A1 *

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WO1995029190A1 (fr) 1995-11-02
JPH10502337A (ja) 1998-03-03
CA2188470A1 (fr) 1995-11-02
FR2719049A1 (fr) 1995-10-27
FR2719049B1 (fr) 1996-06-14
AU2412595A (en) 1995-11-16

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