WO2004014945A1

WO2004014945A1 - Gp41 epitope and uses thereof for the treatment of hiv infections

Info

Publication number: WO2004014945A1
Application number: PCT/IB2003/003149
Authority: WO
Inventors: Lucia Lopalco; Mario Clerici
Original assignee: Istituto Superiore di Sanita ISS; Universita degli Studi di Milano; Ospedale San Raffaele SRL
Current assignee: Istituto Superiore di Sanita ISS; Universita degli Studi di Milano; Ospedale San Raffaele SRL
Priority date: 2002-08-06
Filing date: 2003-08-05
Publication date: 2004-02-19
Anticipated expiration: 2005-02-06
Also published as: ITMI20021781A1; AU2003255866A1; AU2003255866A8

Abstract

Peptides containing a gp41 epitope recognized by IgA of individuals with natural immunity against HIV, antibodies to said peptides and the use thereof in the preventive or therapeutic treatment of HIV infections.

Description

gp41 EPITOPE AND USES THEREOF FOR THE TREATMENT OF HIV INFECTIONS

The present invention regards the identification of a gp41 epitope recognized by the IgA present in the serum and mucosa of individuals with natural immunity to HIV. More specifically the invention is directed to pharmaceutical compositions containing the peptides corresponding to the novel gp41 epitope, the use of said peptides and of antibodies thereto in the immunoprevention or immunotherapy of individuals exposed or at risk of exposure to HIV virus, HIV-infected subjects or AIDS patients.

BACKGROUND OF THE INVENTION

Despite multiple, repeated exposures to HIV some individuals never show any sign of infection (Rowland- Jones SL et al., Curr Opin. Immunol. 1995, 7:448-455; Shearer G et al., Immun. Today 1996, 17 :21-24). A variety of mechanisms are suggested to be associated with this condition including HIV-specific cell mediated and humoral immune responses as well as mutations affecting receptors for virus entry into cells. Numerous reports have documented the presence of HIV-specific cell mediated immunity in HIV- exposed uninfected individuals (EU). Thus, the cell mediated immune responses observed over time in EU include: a) CD4+ and type 1 cytokine- secreting T helper cells; b) CD4+ beta chemokine-producing T helper cells and c) CD8+ and MCH class I-restricted cytotoxic T lymphocytes. HIV- specific humoral immune responses consist in the production of anti cell antibodies and urinary, cervico-vaginal and serum IgA that have the capability of neutralizing primary HIV isolates. Genetic factors can also modulate susceptibility to HIV infection. The first evidence of such factors was provided by the observation that rare individuals homozygous for a deletion (Δ32) within the CCR5 receptor gene are resistant to R5 HIV strains- supported infection. In addition to these mechanisms, exposure to/infection with defective viral strains is postulated as a possible reason explaining the phenomenon of resistance in HIV-exposed individuals. STATE OF THE ART Recently, the identification of a HIV peptide located in the gp41 coiled coil pocket and able to effectively inhibit the entry of HIV- 1 virus, has been reported (Eckert DM et al., Cell. 1999, 99: 103-115; Chan DC et al., Proc Natl Acad Sci 1998: 15613-15617).

Int Arch Allergy Immunol (2000), 121 :80-84, reports a study on the immunogenicity of gpl60-derived peptides and conjugation products thereof. The results indicate that multiple epitopes containing the sequence RILAVERYLKD can induce high levels of epitope-specific antibodies; in addition, the same sequence can induce protective antibodies to HIV (Immunol Today, (1998), 19:586-587). Although the RILAVERYLKD region can induce high humoral specific levels, another region of gp41, having sequence ELDKWA, induces stronger neutralizing humoral responses. The antibody 2F5 produced against the epitope ELDKWA has been the only monoclonal antibody against gp41 to effectively inhibit HIV replication in vitro; therefore 2F5 has been used "in vivo" for the passive immunotherapy of SIV infection in primates(the SIV virus, which is similar to HIV, is primate- specific). Passive immunotherapy with a combination of monoclonal antibodies against gp60 - including 2F5 - proved able to protect primates against mucosal and systemic SIV infection (Nat. Med. 2000; 6:200-206 and 207-210). DESCRIPTION OF THE INVENTION

IgA isolated from sera of HIV-exposed uninfected individuals (EU) and HIV-seropositive individuals (HIV+) were analyzed to establish which region in the HIV envelope is recognized by IgA, and to verify whether different epitopes are responsible for virus neutralization in HIV-resistant and HIV- infected individuals.

Sera from heterosexual couples discordant for HIV serostatus and from HIV-infected subjects were analyzed, using sera from healthy subjects (HC) as negative controls. IgA binding on HIV-env recombinant proteins was assayed. Serum IgA from EU and HIV+ were affinity purified on specific peptides corresponding to different regions of viral envelope, and tested in an HIV neutralization assay using resting and activated PBMC as target cells. Mab 2F5 was used as neutralizing positive control. While IgA of HIV+ patients recognized epitopes expressed in both gpl20 and gp41 env-proteins, IgA of EU exclusively bound epitopes within gp41. Further, in contrast to IgA of HIV+ patients, which recognized the N- terminal portion within the extra-membrane region of gp41, IgA from EU specifically bound a gp41 epitope located in the "leucin-zip" motif in the alpha helix region, corresponding to the peptide with sequence QARILAV (aa 582-588). This peptide ("peptide 1"), as well as the gp41 nona-peptide of sequence LQARILAVE ("peptide 2"), proved particularly effective in terms of immunogenicity and HIV-neutralizing activity. Specifically, antisera from Balb/C mice immunized with peptides 1 and 2 blocked syncytia formation and viral replication. In addition, IgA purified from human sera using peptides 1 and 2 neutralized the infectivity of different primary isolates derived from lymphocytes of HIV-seropositive patients. In particular, peptide-specific IgA inhibited the replication of different viral strains (from different Continents). The dose-response neutralization curve was very similar to that obtained with the monoclonal antibody Mab 2F5.

In a first embodiment, the invention regards the peptides QARILAV (Gin Ala Arg He Leu Ala Val) and LQARILAVE (Leu Gin Ala Arg He Leu Ala Val Glu) isolated from gp41, and their use for the prevention and immunotherapy of HIV-infections. Since the region containing the gp41 epitope is highly conserved in all HIV and lentivirus strains as well, the invention peptides can be used to prevent or treat infections caused by different HIV and lentivirus strains. The peptides can be synthetically prepared, following the procedures described in Merrifield, (1986) Science 232:341-347, and Barany and Merrifield, The Peptides, Gross and Meienhofer, eds (N.Y., Academic Press), pp. 1-284 (1979). The synthesis can be carried out in solution or in solid phase or with an automatic synthesizer (Stewart and Young, Solid Phase Peptide Synthesis, 2^nd ed., Rockford 111., Pierce Chemical Co., 1984). The peptide amino and carboxyl ends can be added with one or more amino acids, or the peptide can be conjugated to different molecules to increase the immunogenicity or improve the pharmacokinetics, provided that these modifications do not cause reduction or loss of the neutralizing activity of the peptide. Examples of such modifications include amidation or esterification of the carboxyl terminus, addition of lipophilic groups (e.g. myristyl group) and glycosylation.

A further embodiment of the invention regards monoclonal or polyclonal antibodies specific for the peptides QARILAV and LQARILAVE. The antibodies can be produced by animal immunization followed by purification, or with genetic engineering techniques, after isolating and identifying the sequence of a specific antibody, preferably a human antibody immunore acting with the invention peptide. Techniques for producing polyclonal antibodies are known in the art and are described for example in Mayer and Walker eds., "Immunochemical Methods in Cell and Molecular Biology", Academic Press London (1987). Methods for purifying antibodies are known in the art and comprise immunoaffinity chromatography. The antibodies may be further manipulated, e.g. through chemical or enzymatic cleavage or by means of genetic engineering, to obtain active fragments such as scFv or Fab. According to a preferred embodiment human antibodies are used, preferably of the IgA type. The antibodies to the invention peptides can be used for passive-immunization of subjects exposed or at risk of exposure to HIV, or for immunotherapy of HIV-infected patients.

In a further embodiment the invention provides pharmaceutical compositions containing an effective amount of at least one of the peptides herein disclosed, or antibodies thereto. The compositions can be administered by the oral, parenteral or topical routes. When peptide-specific IgA are used, the topical route, especially the transmucosal route, is particularly preferred, since the HIV infection is mainly transmitted through the genital mucosa, which is the first physiological barrier to virus entry, and because of IgA blocking of HIV transcytosis from the mucosal to the systemic repertoire. For systemic use, the parenteral administration, e.g. by the intravenous or intramuscular routes, is preferred.

The techniques for preparing pharmaceutical compositions are known to the skilled person and can be found for instance in Remington's Pharmaceutical Science, 17° ed., Mack Publishing Company, Easton, PA (1985), herein incorporated by reference. According to a preferred embodiment, the invention peptides or derivatives thereof are prepared and administered in the form of vaccine. Theory and practice of vaccination are known to anyone skilled in the art, see for example Paul, "Fundamental Immunology" Raven Press, New York (1989) or Cryz, S. J., "Immunotherapy and vaccines", VCH Verlagsgesselshaft (1991). Vaccines are conventionally prepared in the form of injectables, suspensions or solutions, but they can also be used in the form of solid preparations or liposomes. The peptides of the invention, or their derivatives, can be mixed with pharmacologically acceptable excipients, such as emulsifiers, buffering agents and adjuvants which increase the efficacy of the vaccine. The latter can be administered according to single or multiple dosage schedule. Multiple dose provides 1 to 10 separate doses, each containing a quantity of antigen varying from 1 μg to 1000 μg, preferably from 5 to about 250 μg, followed by further doses at subsequent time intervals, necessary to maintain or to reinforce the immune response and, if required, a further dose after several months. In any case the treatment regimen will depend on the response of the treated patient and on his general health conditions.

The immunization with preparations containing an invention peptide or derivatives thereof can be applied to subjects exposed or at risk of exposure to HIV virus or as a therapeutic support for HIV-infected patients.

The invention is also directed to a nucleic acid molecule encoding an invention peptide. The sequences encoding peptides 1 and 2 are preferably CAGGCAAGAATCCTGGCTGTG and CTCCAGGCAAGAATCCTGGCTGTGGAA, respectively. The choice of the coding nucleotide sequence depends on the expression system to be used, and can be varied by virtue of genetic code degeneracy.

The invention further comprises an expression vector wherein said nucleic acid sequence is operatively linked to a heterologous promoter, and a host cell for peptide production containing said expression vector.

The DNA molecules coding for the invention peptides and their genetic constructs can be used in the vaccination of subjects exposed or at risk of exposure to HIV, or in the treatment of HIV-infected patients. The DNA immunization can be carried out according to known procedures (Donnelly J.J. et al., 1994, The immunologist 2: 1). The intramuscular administration route is preferred, but the parenteral and mucosal routes can be used as well (PNAS 1986, 83, 9551; WO90/11092). In addition, the DNA can be adsorbed onto gold particles and transdermally administered with a biolistic apparatus (Johnston, 1992 Nature, 356, 152).

The peptide-specific amino acid and nucleotide sequences can be used for the development of a prognostic test. Accordingly, the peptide may represent a marker of previously-occurred exposure without viral transmission. This prognostic test could be applied to any population at risk of exposure to HIV, so as to exactly determine the number and type of individuals that should be subjected to therapeutic prophylaxis.

DESCRIPTION OF THE FIGURES

Fig. 1 Neutralizing activity of i) total IgA from EU and HIV+, ii) purified fraction containing anti-peptide 1 and 2 (pep#l/2) IgA from EU, and iii) fraction without anti-peptide 1/2 IgA from HIV+; total IgA from HC

(healthy control) and the monoclonal antibody 2F5 were used as negative and positive controls, respectively.

Fig. 2 Binding to gp41 of mouse antisera against peptides 1 and 2. A non-correlated mouse serum was used as negative control.

Fig. 3 Modulation of HIV-1 infectivity by mouse antisera against peptides 1-2. A non-correlated mouse serum was used as negative control.

Fig. 4 HIV infection through the mucosa (transcytosis)

Fig. 5 In vitro model of transcytosis inhibition. Fig. 6 Transcytosis inhibition by IgA specific for the sequence

QUARILAV and by total IgA from either EU or negative controls.

Fig. 7 Titration of sera from mice immunized with Leu581 peptide, corresponding to the III, V, VII and XI immunization.

Fig. 8 Titration of sera from mice immunized with peptide QUARILAV, corresponding to immunization III, V, VII and XI.

Fig. 9 Quantitative analysis of total antibodies and antibodies specific for different peptides after immunizations of two different mice.

Fig. 10 Syncytia formation inhibition by different concentrations of specific antibodies against the two peptides. MATERIALS AND METHODS

IgA purification

Sepharose 4B CNBr activated (Pharmacia, Uppsala, Sweden) was coupled with rabbit anti human IgA (2mg/μl) (Aldrich, Milan, Italy) by standard methods. In brief, 96 μl of serum were incubated at room temperature for 15' on columns containing 2400 μl of sepharose -anti IgA. The columns were washed, IgA were eluted with Glycine/NaCl 0.2 M, and the solution was neutralized with 1M Tris pH 11. Purified IgA were concentrated on Ultrafree- 15 Biomax30 membranes (cut off 30kDa)(Milliρore, Bedford, MA, USA). IgA were also purified on sheep anti human IgA sepharose-conjugated columns to exclude the presence of cross-reactive material using the same procedure.

Binding of peptides to dynabeads

HIV peptides were directly coated onto Dynabeads M280 tosylactivated (Dynal, Oslo, Norway) by standard procedures. Briefly, 3x10^ beads were incubated with 9 μg of each peptide in borate buffer pH 9.5 for 16 h at 37°C. After 4 washes (in PBS), the peptide-conjugated beads were ready for use.

Binding and elution of IgA to peptides conjugated to dynabeads

Binding of IgA from EU, healthy donors and HIV-seropositive subjects to each env peptide/bead was obtained by incubating 9 μg of purified IgA to 9 μg peptide/bead for lh at 4°C. Beads-conjugated IgA were eluted from the beads in acetic acid 0.5 M and dialized.

Binding of IgA to recombinant env proteins

In order to determine the concentration of IgA eluted from each peptide/bead complex, the fractions contaning IgA/peptide enriched and

IgA/peptide depleted, were tested in ELISA assay. Microwells plates were coated with all fractions at different dilutions (starting from 1/1 to 1/16 by two-fold dilutions). IgA binding was revealed with HRP conjugated rabbit anti human IgA (Dako, Santa Barbara, CA).

The IgA eluted from each peptide/bead complex were tested in ELISA (Table). Microwells plates were coated with gp41/gpl20 recombinant proteins at 0.1 μg/wells by overnight incubation in NaHC0₃/Na₂C0₃ buffer. The plates were satured for lh with PBS and 3% BSA. Eluted IgA were added and incubated for lh at 37°C. IgA binding was revealed with HRP conjugated rabbit anti human IgA (Dako, Santa Barbara, CA). The enzymatic reaction was developed and read at 492nm. Competition assays were performed by preincubating IgA (1.5 nM) with 100 fold in excess of specific synthetic peptides (150 nM) for 30 min at 37°C prior to addition to the microplates.

To perform Western blots of gpl20 recombinant protein, 30 μg of protein were loaded on 10% polyacrylamide gel, run, and transferred to nitrocellulose membranes; a 3 hours incubation with 50 μg of purified IgA from either EU; HIV+; or healthy controls followed these steps.The membranes were then incubated for lh with a 1/1000 dilution of a peroxidase conjugated rabbit antihuman IgA (Dako), and revealed with the ECL Western blotting system (Amersham, Buchinghamshire, England).

Generation of mouse serum against peptides 1 and 2 from gp41

BALB/c mice were immunized i.p. every 15 days with peptides QARILAV or LQARILAVE (three mice per peptide) or with an unrelated control peptide (VQGEESNDK) (two mice); both peptides had previously been conjugated to KLH (50 μg/dose). Serum was monitored for the presence of antibodies against the immunogens. When the specific serum titer was > 1 :

1,500 dilution, mice were sacrificed and sera were tested in neutralization assay.

Virus isolation and characterization

HIV was isolated from PBMC of HIV-seropositive individuals by co- coltivation with PHA- stimulated PBMC from two healthy donors. The cultures were maintained as long as increasing levels of HIV-p24 antigen were detectable in two consecutive determinations (ranging from 10 to 18 days). The infectivity (LD50) of each virus isolate was determined on PBMC from a single donor. Virus was diluted by five-fold dilutions, beginning with 1/5. Each dilution (150 μl) was added to six parallel wells of a round-bottomed microtiter plate (Nunc, Roskilde, Denmark) containing 10^ resting or activated PBMC in 75 μl of medium, incubated for two hours, washed and resuspended in medium additioned of PHA (Sigma) and lOU/ml rIL2

(Amersham). After 7-9 days samples were analysed for HIV-1 p24 antigen. ID 50 titers were defined as the reciprocal of the virus dilution resulting in

50% positive wells (Reed-Muench calculation). All isolates were tested on

U87 transfected cell line to determine the coreceptor usage of each primary virus (strains R5 and R5/X4/R3).

Virus neutralization assays Neutralization activity of purified IgA was measured by standardized methods. Briefly, 2x10 resting or activated (in this latter case, the cells were cultured in medium containing PHA and IL2 for 48h before the neutralizing assay) PBMC from a healthy donor were added to 96 microplate wells. Seventy-five μl of primary virus dilutions containing TCID50 (range 10 to 32) of each HIV-1 primary isolate was added to 75 μl of serial dilutions of serum purified IgA fraction (30, 10, 3, 1 μg/ml) and serum IgA depleted fraction, incubated for lh and added to the PBMC. Cultures were then incubated for 2 hours, washed and resuspended in PHA and IL2-containing medium. HIV-p24 levels were determined in the supernatants 7, 9, and 11 days after infection. Negative (purified IgA of healthy donors) and positive (IgA from HIV+ patients and gp41 specific monoclonal antibody 2F5) controls were run in each assay and each sample was tested in duplicate. The mean value obtained with each sample was compared with the mean values of the six corresponding replicates without the addition of antibodies and expressed as percentage of inhibition. O.D. values in the absence of IgA ranged from 1.55 to 1.83. Syncytia inhibition assay

8E5 (LAV) cells and MOLT3 cells were grown in RPMI 1640 at lO^/ml, washed, mixed in 1:2 proportion, and distributed in V-shaped microtiter wells (1.6x10^ cells/well) together with IgA from EU or from healthy controls or mouse antisera in appropriate dilutions. After 30 min incubation at 37°C, the cells were sedimented by centrifugation and incubated as a pellet for 3h at 37°C. Syncytia (10 to more than 100 cells) were scored with an inverted microscope after transfer of the pellet to flat-bottom well microtiter plates. In uninhibited control samples, 100-130 large syncytia per well were counted. Competition of IgA inhibitory effect by specific peptides was tested by preincubating IgA at 200 μg/ml with each peptide (200 μg/ml) for 30 min at 37°C prior to addition to the microplates. RESULTS

Identification of the epitope recognized by HIV- 1 -specific human IgA

IgA purified from EU or HIV+ patients were coupled to dynabeads previously conjugated to peptides corresponding to the proteins of the HIV envelope. Specific binding to gp41 was observed with IgA from EU. IgA coupled to the different peptides were eluted and utilized in gp41 binding assays at a concentration of 4 μg/ml. When IgA of EU were analyzed, specific binding to gp41 was observed only when antibodies eluted from peptide 1/2- coated beads were employed (Table). In contrast, when IgA obtained from

HIV-infected individuals were analyzed, gp41 binding was observed with antibodies eluted from peptide# 13 -coated beads (Table). Total IgA samples from 2 healthy donors were pooled and used as negative controls (Table).

Table. ELISA reactivity on gp41 recombinant protein by human IgA coupled to different peptides. Peptide pep#13 (NYTSLIHSLIEESQNQQEKNEQELLELDKWAS) corresponds to the gp41 region recognized by the monoclonal antibody 2F5. The results are expressed as O.D. value (optical density) obtained with samples from 6 EU, 12 HIV+ and 6 HC.

Affinity-purified anti-peptide 1/2 human IgA inhibit HIV- 1 replication To characterize the biologic activity of IgA specific for QARILAV and LQARILAVE peptides, IgA bound to these same peptides were eluted and evaluated in neutralization assay. Peptide-specific human IgA, affmity- purified from sera, were tested in HIV neutralization assay using the primary isolates of HIV. The neutralizing titers obtained with IgA eluted from peptides 1/2 were higher (IC₅₀ of 1.8 μg/ml) than those of total IgA (IC₅₀ of 10 μg/ml). The IC₅₀ obtained with total IgA was of > 100 μg/ml (Figure 1).

To confirm that purified anti-peptide 1/2 human IgA are responsible for the HIV inhibitory effect, a neutralization assay using activated PBMC as target cells was used. In this assay, total IgA from representative EU, HC and HIV+ samples, as well as peptidel/2 specific EU IgA purified fraction and pepl/2-IgA depleted fraction from both EU and HIV+ subjects were utilized. A neutralizing anti gp41 monoclonal antibody (2F5) (Muster T. et al., J Vitol. 1994, 68:4031-4034) was used as positive control. A strong neutralizing activity was observed when: i) total IgA from both EU and HIV+; ii) pep 1/2- IgA purified fraction from EU; or iii) pepl/2-IgA-depleted fraction from HIV+ containing specific IgA against gp41 regions different from the region (L)QARILAV(E), were utilized in the assay. Interestingly, the dose dipendent neutralizization behaviour of peptidel/2 specific IgA enriched fraction is very similar to that obtained with Mab 2F5. The IC₅₀ obtained with total EU IgA was 0.51 μg/ml while IC₅₀ obtained with peptides 1 and 2 specific IgA was 0.17 μg/ml. Similar results were obtained in resting cell assays.

Peptides 1- and 2-specifϊc mouse sera efficiently neutralize HIV-1 primary isolates

BALB/c mice were immunized with either 50 μg of peptides 1 or 2 or a control - peptide. The binding to gp41 by the corresponding antisera was evaluated with ELISA assay. A dose-dependent binding specific for all the three sera was observed (Fig. 2). The specific sera were then tested in a neutralization assay where activated PBMC cells were infected with the HIV-1 primary isolate

HIV#40 (Figure 3) in the presence of four different antiserum dilutions (ranging from 1/40 to 1/1080). As shown in Figure 3, the specific mouse antisera inhibited HIV replication in a dose-dependent manner. No inhibition was observed using sera from mice immunized with the control peptide.

Antibodies to peptides 1 and 2 inhibit HIV- 1 viral entry

Mouse antisera to peptides 1 and 2 effectively inhibit viral replication.

We thus tested whether these antibodies would interfere with viral entry by analyzing the ability of IgA purified from sera of EU and HC and sera of mice immunized either with peptides 1 or 2 or the control peptide to inhibit syncytium induction. Results showed that IgA from EU and sera of mice immunized with peptides 1 or 2 greatly reduced syncytium induction. In contrast, IgA of HC and sera of mice immunized with the control peptide had no effect. Pre-incubation of the mouse antisera with peptides 1 or 2 but not with pep#13 inhibited the blocking capacity.

IgA assays specific for QARILAV

IgA specific for QARILAV from the vaginal fluids of two EU were assayed in an in vitro model mimicking the in vivo pattern of HIV transmission through the genital mucosa, whereby the IgA should block the passage of HIV virus from the mucosal to the systemic repertoire. Since the studied EU had multiple sexual intercourses with HIV-positive subjects, the analysis of their mucosal system is of particular relevance.

Figures 4 and 5 illustrate an in vivo and, respectively, in vitro- reproduced mucosa model.

Figure 6 shows the in vitro HIV inhibition by QARILAV-specific IgA. Balb/C mouse immunization with QARILA V and with QARILA VERYLKDQQLLG

To verify whether the humoral responses identified in EU could be reproduced in animal models, Balb/C mice were immunized through the intraperitoneal route with the peptide QARILAV, or with QARILA VERYLKDQQLLG (leu 581) peptide, already described in the literature as immunogenic (i.e. inducing specific antibodies). The QARILAV peptide immunogenicity and ability to induce neutralizing antibodies was evaluated in comparison with leu 581. Eleven immunizations were performed with intervals of 10 days and hyperimmune sera were analysed either in peptide-binding assays (to determine the immunogenicity) or in viral infection inhibition assays (syncytia formation inhibition assay).

Figures 7 and 8 show the titration curves of the peptide-specific antibodies against Leu 581 and QARILAV, respectively, after several immunizations.

Figure 9 reports the concentrations of total antibodies and of the antibodies specific for the peptides leu 581 and QARILAV. Serum immunoglobulins from non-hyperimmune mouse were used as control. As shown in Figure 2, the QARILAV peptide is able to induce a stronger immune response as compared with that induced by Leu 581 (12% and 9% Ig specific for QARILAV and for Leu581, respectively). Viral infection inhibition test

To verify the inhibitory effect of the immunoglobulins specific for the two peptides, a viral infection inhibition test for the quantification of HIV- induced syncytia formation has been utilized. As shown in Figure 10, the immunoglobulins specific for the QARILAV peptide possess a higher syncytia-formation inhibitory activity than that of antibodies specific for the Leu581 peptide.

Claims

I. Immunogenic peptide isolated from gp41, having amino acid sequence selected from QARILAV and LQARILAVE.

2. Nucleic acid molecule encoding the peptide of claim 1.

3. Nucleic acid molecule according to claim 2, having nucleotide sequence CTCCAGGCAAGAATCCTGGCTGTGGAA or CAGGCAAGAATCCTGGCTGTG.

4. Monoclonal or polyclonal antibody to the peptide of claim 1.

5. Antibody according to claim 4, of the IgA type.

6. Pharmaceutical composition containing an active ingredient selected from the peptide according to claim 1, the antibody according to claims 4-5, the nucleic acid molecule according to claims 2-3.

7. Pharmaceutical composition according to claim 6, containing the peptide according to claim 1 or the nucleic acid molecule according to claims

3-4, in the form of vaccine.

8. Pharmaceutical composition according to claim 6, containing an active ingredient selected from the peptide according to claim 1, the antibody according to claims 4-5, the nucleic acid molecule according to claims 2-3, wherein said pharmaceutical composition is suitable for oral administration.

9. Pharmaceutical composition according to claim 6, containing the antibody according to claims 4-5, wherein said pharmaceutical form is suitable for topical administration.

10. Use of the peptide of claim 1, of the antibody according to claims 4-5 or of a nucleic acid molecule according to claims 2-3 for the preparation of a medicament useful for the preventive or therapeutic treatment of HIV infection.

I I. Use according to claim 10, wherein said medicament is in form of vaccine.

12. Use according to claim 10, wherein said medicament is suitable for oral administration.

13. Use according to claim 10 of the antibody according to claims 4-5, wherein said medicament is suitable for topical administration.

14. Use according to claims 10-13, for the preventive or therapeutic treatment of infections caused by HIV-1 or other lentivirus.