EP0590057A1

EP0590057A1 - MIMIC PEPTIDES OF gp120

Info

Publication number: EP0590057A1
Application number: EP92914211A
Authority: EP
Inventors: Malcolm L. Gefter; Kenneth L. Melmon
Original assignee: Immulogic Pharmaceutical Corp
Current assignee: Immulogic Pharmaceutical Corp
Priority date: 1991-06-19
Filing date: 1992-06-19
Publication date: 1994-04-06
Also published as: CA2111681A1; WO1992022579A1

Abstract

Cette invention concerne des peptides comprenant une séquence primaire d'aides aminés qui n'est pas liée à la gp120. Les peptides se caractérisent par leur aptitude à stimuler la production d'anticorps anti-gp120, plus particulièrement d'anticorps anti-boucle-V3 lorsqu'on les administre à un mammifère. Ces peptides se caractérisent également par leur capacité à entraver avec la fonction virale. On peut formuler lesdits peptides sous forme d'une composition vaccinale et les administrer pour stimuler la réponse immunitaire neutralisant le VIH-1.This invention relates to peptides comprising a primary amino helper sequence that is unrelated to gp120. The peptides are characterized by their ability to stimulate the production of anti-gp120 antibodies, more particularly of anti-loop-V3 antibodies when they are administered to a mammal. These peptides are also characterized by their ability to interfere with viral function. Said peptides can be formulated as a vaccine composition and administered to stimulate the HIV-1 neutralizing immune response.

Description

MIMIC PEPTIDES OF σpl20

Background of the Invention

Human immunodeficiency virus (HIV) is the cause of acquired immunodeficiency syndrome and the HIV external envelope glycoprotein, gpl20/ is associated with viral infectivity and cytopathology including cell fusion. gpl20 binds the cellular receptor of the virus, CD4, and cells expressing the envelope fuse with CD4-positive cells in culture. Fusion of infected and noninfected cells and infection by cell-free virions are important routes of HIV infection.

It has been reported by Robey et al. (Proc. Natl. Acad. Sci. USA & :7023-7027 (1986)) that gpl20 purified from virus-infected cells elicits antibodies that neutralize the infectivity of HIV. Recombinant gpl20 or gplβO expressed in a variety of cell types have also been shown to elicit neutralizing antibodies (see e.g., Lasky et al.. Science 221:209-233 (1986); Rusche et al., Proc. Natl. Acad. Sci. USA 84;1-5 (1987); and Steimer et al.. Vaccines 87:236-241 (1987)). The amino acid sequence of the envelope is known to vary between different HIV isolates and the neutralizing antibodies elicited by the envelope of one HIV isolate are usually effective against only a subset of heterologous isolates. This observation suggests that such neutralizing antibodies are directed to a variable region of the envelope. This idea is supported by the fact that observation suggests that such neutralizing antibodies are directed to a variable region of the envelope. This idea is supported by the fact that neutralizing antibodies are elicited by an E. coli-produced recombinant fragment from the carboxyl-terminal region of gpl20, PB1, that contains 37% of gpl20 and is from a variable region of the envelope. This fragment contains the dominant neutralizing epitopes (epitope being the basic element or smallest unit of recognition by a receptor such as an antibody), and a more complete understanding of the number, the location, and the potential role of antibodies to these epitopes in preventing viral infection may facilitate development of a subunit vaccine able to induce immunity to diverse HIV isolates.

Matsushita et al. tJ. Virol. 62:2107-2114 (1988)) reported the identification of a monoclonal antibody which is characterized by the ability to neutralize infection by cell-free virus, and the ability to prevent fusion of virus-infected cells. The antibody binding site was mapped to a 25-amino acid segment of gpl20, referred to herein as the V3-loop. This region is known to contain major neutralizing epitopes. Peptides which stimulate the production of antibodies which are reactive with these immunodominant epitopes yet which are not found in the primary sequence of the gpl20 protein would be useful for immunization, for example. Such peptides are referred to herein as mimic peptides.

Summary of the Invention

This invention relates to peptides having a primary amino acid sequence which is unrelated to the primary gpl20 amino acid sequence and are, by themselves, or conjugated to an immunogenic carrier molecule, able to stimulate the production of anti-gpl20 antibodies. The anti-gpl20 antibodies produced in response to peptides of the present invention are directed toward a region of the gpl20 molecule known as the V3-loop, which is known to contain important epitopes.

The peptides can be formulated in a physiologically acceptable carrier for use as a vaccine composition. This composition can optionally include an adjuvant. In addition, the peptide can be conjugated to an immunogenic carrier molecule to increase immunogenicity. In this context the peptide serve as an immunogen.

The invention also pertains to peptides which serve as antiviral agents and a method for interfering with viral propagation in an individual. The method involves administering an effective amount of a therapeutic composition comprising a peptide in a physiological carrier. The peptide has a primary amino acid sequence which is unrelated to the primary amino acid sequence of an essential protein of a virus (e.g., gpl20) and the peptide has the ability to interfere with a component involved in the function of the virus. In this context, the peptide serves as an antiviral agent.

In another aspect, the invention pertains to a method for screening an epitope library with a non-isolate specific antibody. Such antibodies typically do not bind antigen with high affinity. Brief Description of the Drawings

Figure 1 is a schematic representation of results from a competition assay for V3-loop binding.

Figure 2 is a schematic representation of results from a competition assay for recombinant gpl20 binding.

Figure 3 is a schematic representation of results from a direct binding assay with serum from mice immunized with M306 G3. Figure 4 is a schematic representation of results from a direct binding assay with serum from mice immunized with M306 G3 conjugated to an immunogenic carrier molecule.

Figure 5 is a schematic representation of results from a competition experiment in which V3-loop was attached to an ELISA plate and reacted with serum from mice immunized with M306 G3 conjugated to an immunogenic carrier molecule mixed with various peptides. Figure 6 is a schematic representation of results from a competition experiment in which M306 G3 conjugated to bovine serum albumin was fixed to an ELISA plate and reacted with serum from mice immunized with M306 G3 and conjugated to an immunogenic carrier molecule mixed with various peptides.

Figure 7 is a schematic representation of results from a competition experiment in which V3-loop was fixed to an ELISA plate and the antibody was an affinity purified rat monoclonal known to be HIV neutralizing. Figure 8 is a schematic representation of results from a competition experiment in which recombinant gpl20 was fixed to an ELISA plate and the antibody was an affinity purified rat monoclonal known to be HIV neutralizing.

Detailed Description of the Invention

One embodiment of subject invention is based on Applicants' discovery of peptides, unrelated in primary amino acid sequence to the primary amino acid sequence of gpl20, which stimulate the production of anti-gpl20 antibodies when administered in vivo to a mammal. The primary amino acid sequence refers to the linear order of amino acid residues in a peptide or protein. Peptides of the invention were identified initially by screening an epitope library with a rat monoclonal antibody known to be HIV neutralizing in an in vitro assay. Such a method is described, for example, by Scott and Smith (Science 24_iJ386-390 (1990)). Briefly, an epitope library is a vast mixture of filamentous phage clones, each displaying one peptide sequence on the virion surface. The survey is accomplished using a binding protein (e.g., a monoclonal antibody) to affinity-purify phage that display tightly-binding peptides and propagating the purified phage in IL. coli. The amino acid sequence of the peptides displayed on the phage are then determined by sequencing the corresponding coding region in the viral DNAs. Peptides identified in this manner that are also capable of competing for antibody binding with the natural epitope are referred to herein as mimic peptides because they mimic the natural epitope, but in most instances are unrelated in primary sequence to the natural epitope.

More specifically, the steps involved in the screening of an epitope library include providing an epitope library containing bacterial cells infected with fusion phage. The fusion phage are then contacted with a biotinylated antibody under conditions appropriate for the binding of the antibody to peptides displayed on the surface of the bacterial cells infected by the fusion phage. The mixture is then contacted with a streptavidin-coated substrate under conditions appropriate for the binding of biotin to streptavidin. Non-specifically bound phage are removed with a wash step. Bound phage are eluted and used to infect I coli cells and peptide sequences binding to the rat monoclonal antibody were identified. Although these peptide sequences are fourteen amino acid residues in length, it is believed that some or all of the hexapeptide amino acid residues 5 through 10 are essential for the binding of the peptides to the rat monoclonal antibody since amino acid residues 1 through 4 and 11 through 14 were identical for all the peptides in the epitope library. However, it is possible that some or all of the amino acid residues 1 through 4 and 11 through 14 are critical since amino acid residues 1 through 4 and 11 through 14 were part of the peptide that actually demonstrated function in the Exemplification. Peptides having the same sequences as the peptides identified by screening the epitope library, except for the addition of amino acid 15 (alanine) were synthesized and assayed for the ability to stimulate an anti-gpl20 or anti-V3-loop response when administered to mice in vivo, as described in the Exemplification below. Murine antisera was tested for reactivity in a direct binding assay as described in detail in the Exemplification section. Three peptides were identified which stimulate a response which is cross-reactive with the natural HIV epitope. The three peptides are 15 amino acid residues in length and can be represented generically as follows:

Ala Asp Gly Ala AAl Gin AA2 Asn Arg AA3 Gly Ala Ala Gly Ala,

In the generic formula above the symbols AAl, AA2 and AA3 represent amino acid residues which are not perfectly conserved in the three mimic peptides described herein. Thus, the peptides of this invention include peptides of the generic formula set forth above wherein AAl, AA2 and AA3 represent any amino acid residue. In addition, the peptides of the invention include fragments of the 15 amino acid sequence which have the desired properties discussed herein. More preferably, AAl and AA3 are selected from the group of amino acids consisting of serine, threonine, methionine and cysteine and AA2 is selected from the group consisting of lysine, arginine, histidine, serine and threonine. Due to the critical nature of the hexapeptide, the peptides of this invention can also be represented generically including the following sequence: AA1 Gin AA2 Asn Arg AA3

where symbols AAl, AA2 and AA3 are the same as above described, the preferred peptides including the following sequences being as follows:

Ser Gin Thr Asn Arg Met;

Met Gin Ser Asn Arg Ser; and Ser Gin Arg Asn Arg Ser.

Peptides of the generic formula set forth above can be routinely synthesized by standard chemical techniques or by recombinant DNA methodology. Such techniques are well known in the art. Peptide synthesizers are available commercially and standard techniques have been described in numerous publications including, for example, Merrifield (J. Chem. Soc. 85:2149-2154 (1963)) and Hunkapillar et al. (Nature 310:105-111 (1984)). It is well known to one skilled in the art that the immunogenicity of small non-, or weakly-immunogenic molecules (haptens) can be increased by conjugating the weakly- immunogenic molecule to a carrier protein. Commonly used carrier proteins include keyhole limpet hemacyanin and bovine serum albumin. The peptides of this invention can be conjugated to a carrier molecule, using conventional methods, to enhance their immunogenic properties.

Another embodiment of the subject invention is based on Applicants' discovery of peptides, unrelated in primary amino acid sequence to the primary amino acid sequence of an essential protein of a virus (e.g., HIV gpl20), which have the ability to interfere with a component involved in the function of the virus. In a preferred embodiment, these peptides have the ability by themselves, or conjugated to an immunogenic carrier molecule, to stimulate the production of antibodies specifically reactive with the essential viral protein when administered to a mammal such as the peptides set forth above which are mimic peptides for the V3 loop. One skilled in the art will recognize that the amino acid sequences of the peptides set forth above represent only the essential portion required for immunogenic or anti-viral activity. A peptide which is useful as an immunogen or anti-viral agent is not necessarily limited to a minimal sequence shown above, but must have at least a portion of an amino acid sequence shown above which provides immunogenic or anti-viral activity. In addition, peptides which include modified amino acids are included within the scope of the invention provided that the modifications (e.g., conservative substitutions) do not adversely affect the functional characteristics of the peptides.

Uses for the mimic peptides

It is well known in the art that antibodies directed to the V3-loop can prevent viral infection by both free and cell-associated virus. The mimic peptides can be used, for example, as immunogens to stimulate an anti-gρl20 immune response in an individual. Such a response could be protective in the sense that the individual to whom the peptides are administered may be immune to infection by the HIV virus. To be used as a vaccine, the mimic peptides are formulated in a composition and administered according to conventional protocols. Such a composition includes at least one mimic peptide, by itself or conjugated to an immunogenic carrier molecule, in an amount sufficient to stimulate an immune response in a host. The mimic peptide is contained in a physiologically acceptable carrier. Such carriers include, for example, fillers, non-toxic buffers, physiological saline solution, etc. The composition can also include adjuvants, protease inhibitors, lymphokines, or compatible drugs where such a combination is seen as desirable or advantageous. Although the role of the V3-loop has not been elucidated, it appears that the V3-loop has a viral function which leads to viral propagation. Other pathogenic viruses contain proteins which have a viral function which lead to viral propagation. Peptides which are unrelated in primary amino acid sequence to the primary amino acid sequence of an essential protein of a virus, preferably mimic peptides, can therefore be used in a therapeutic composition to interfere with viral replication in an individual. This can occur, for example, by interference of the binding of a viral product with a cellular or viral component. The mimic peptides can be used as decoy molecules to inhibit the natural function of an essential viral protein (e.g., the HIV gpl20 protein) . By introducing an effective amount of the mimic peptides to an infected or non-infected individual, it is reasonable to predict that their presence will interfere with viral replication thereby slowing or preventing the spread of the infection.

EXEMPLIFICATION

Direct binding assay

Direct binding between test peptides and test antibody (affinity purified or in serum) was determined by ELISA assay. Costar ELA/RIA plates were coated by overnight incubation with 50 μl of a solution containing the test peptides or controls. Following the overnight incubation, the plates were washed 3 times with 1 X Phosphate Buffered Saline (PBS) . The plates were then blocked with 1% Bovine Serum Albumin (BSA) essentially globulin free (Sigma Cat. #7638) at room temperature for about 1 hour or more.

The blocker was removed and about 100 μl of affinity purified 1° antibody (or test serum) was added in a dilution series. The dilutions were made in PBS-T/FCS (1 X PBS, 0.05% Tween-20, 2.5% Fetal Calf Serum (FCS) . The serum was incubated on the plate for about 1 hour at room temperature. Following this incubation, the plates were washed 3 times with PBS-T/FCS. 2° antibody was then added (at about 100 μl per well) and incubated at room temperature for about 1 hour. The 2° antibody is typically an anti-species antibody which is biotin labeled. This incubation is followed by 3 PBS-T/FCS washes. A Streptavidin-HRP (horseradish peroxidase) conjugate (Southern Biotechnology Associates, Birmingham, AL) was added at an appropriate dilution (typically 1/10,000) in PBS-T/FCS and incubated at room temperature for about 1 hour. This step was followed by 3 washes with PBS-T/FCS. The assay was developed using the TMB Microwell Substrate System (Kirkegaard & Perry Labs, Gaithersburg, MD) . The optical density (OD) was read on an ELISA reader at 450 nanometers.

Competition experiments

Competition experiments were carried out by mixing each test peptide and control peptide in solution with a test antibody and determining the effect (if any) of the peptide in solution on binding of the antibody to a peptide attached to the plate in an ELISA assay. The protocol is essentially as described above in connection with the ELISA assay except that peptides (or peptide conjugates) to be tested for ability to compete with the fixed peptides are mixed with the antibody and incubated for about 30 minutes prior to the addition of the antibody to the ELISA plate.

Identification of antibodies reactive with V3-1OOP Four rat monoclonal antibodies which react specifically with HIV-IIIB were tested for V3-loop binding. Each of these antibodies exhibits anti-syncytia activity in an in vitro assay. The antibodies were designated 24D3, 20G3, 1G7 and 28G12. In a direct binding assay, of the type described above, polystyrene plates were coated with cyclized

V3-loop (50 μg/ml), non-cyclized V3-loop (50 μg/ml) and recombinant gpl20 (2 μg/ml) . Each of the 4 rat monoclonal antibodies was tested in the assay. In addition a murine control which was known to bind V3-loop was included as a positive control. The results from this binding assay clearly showed that all four rat monoclonal antibodies react specifically with the cyclized and non-cyclized form of V3-loop. Antibody 1G7 demonstrated the strongest reactivity.

Two of the antibodies were biotinylated (1G7 and 28G12) by conventional methods. Specifically, in separate 1.5 ml polypropylene tubes, 167 μl (500 μg) of 28G12 and 250 μl of 1G7 (500 μg) were added. To each tube was added 12 μl of a biotin solution and the tubes were incubated at room temperature, with agitation, for 2 hours. The biotin solution was prepared by dissolving 1-3 mgs NHS-LC Biotin (Pierce Cat. #21335; MW 556.59 daltons) in 456.6 μl distilled water/mg of reagents. The final concentration was 3.93 mM. The solution was made fresh each time the experiment was performed.

To prevent biotinylation of the carrier ovalbumin, 500 μl of IM ethanalamine was added (pH 9) and the mixture was incubated at room temperature for 2 hours. 20 μl of carrier ovalbumin (10 μg/ml was also added) and the biotinylated antibodies were diluted against 1 X TBS overnight. The concentration of each was determined to be about 400-500 μg/ml. The material was concentrated in a Centricon^R 30-kilodalton ultrafilter (Amicon), and washed three times with 2ml TBS to remove unconjugated biotin. 300 μl Of TBS/0.02% NaN3 and 40 μl (7.44 X 10¹² virions) UV-killed blocking phage was added directly to the Centricon^R. The mixture was vortexed to thoroughly mix the blocking phage with the antibody solution, then reconcentrated. The blocking phage are meant to block antibodies that cross-react with the filamentous phage in general, apart from the foreign peptide they display. The retentate was collected in the conical cup by back-centrifugation, as described in the Centricon^R instructions, and stored at 4°C.

Screening of the epitope library

Biotinylated antibodies, prepared as described above, were used to screen an epitope library (Scott and Smith, Science 249:386-390 (1990)). The epitope library was screened essentially as described by Scott and Smith (supra) . In this way, peptides were discovered which specifically bind the HIV neutralizing rat monoclonal antibodies described above. Four 15 amino acid length synthetic peptides were produced which contain the 14 amino acid peptide identified by screening the epitope library. These peptides are referred to herein as M303 G7, M305 G14, M306 G3 and M307 G10 and the sequences of these peptides are as follows:

Ser Gly Ala Ala Gly Ala For purposes of reference, the V3-loop sequence is: Gly Cys Thr Arg Pro Asn Asn Asn Thr Arg Lys Ser lie Arg lie Gin Arg Gly Pro Gly Arg Ala Phe Val Thr lie Gly Lys lie Gly Asn Met Arg Gin Ala His Cys Gly. The cyclized form of the V3-loop is generated by joining the amino terminus and the carboxy terminus of the linear molecule.

Reactivity of M303 G7. M305 G14 and M306 G3 with rat monoclonal 28GI2 The M303 G7, M305 G14, M306 G3 and M307 G10 peptides were synthesized by conventional methods and tested for direct binding with rat monoclonal antibody 28G12 by the ELISA assay described above. Each of the four peptides was attached to separate wells of an ELISA plate. Appropriate controls were included such as an V3-loop positive control in an ELISA well and a BSA negative control in a separate well.

In the direct binding assay, the antibody showed strong binding to the positive controls (V3-loop and recombinant gpl20) but showed binding to M303 G7, M305 G14, M306 G3 and M307 G10 at near background levels or slightly elevated. This may be an artifact of the small size of these peptides (15 amino acids as compared to 38 for V3-loop) .

Surprising results were observed when the test peptides were tested in the competition assay described above. Specifically, when three of the four test peptides were incubated with the 28G12 antibody in solution prior to incubation with plates coated with V3-loop or recombinant gpl20 a significant decrease in antibody binding, as evidenced by a decrease on the OD45₀ determination, was observed. When tested in this manner, the peptide M307 G10 was the only test peptide that did not result in a significant decrease in antibody binding. This M307 G10 peptide had glycine as the ninth amino acid residue, whereas the other three test peptides, M303 G7, M305 G14 and M306 G3 all had arginine as the ninth amino acid residue. In the competition experiment, the control peptides used were lambda repressor amino acid residues 12-26, said peptide having the sequence Leu Glu Asp Ala Arg Arg Leu Lys Ala lie Tyr Glu Lys Lys Lys (CONTROL in Figures 1-7) and a synthesized peptide, M304 P8, containing the sequence of a 14 amino acid peptide that did not bind biotinylated antibody during the screening of the epitope library, said M304 P8 peptide having the sequence Ala Asp Gly Ala lie Ser Asn Leu lie Ser Gly Ala Ala Gly Ala. The results of this experiment are shown in Figures 1 and 2.

immunization of mice with mimic peptides

BALB/C mice were immunized using conventional methods with each of the three mimic peptides. The mice were primed on Day 0 with 50 μg per mouse of either M306 G3 in complete Freund's adjuvant (CFA) or M306 G3 conjugated to keyhole limpet hemacyanin (KLH) in CFA. At 21 days post-immunization, the mice were tail bled for a 1° response and the serum was collected. The mice were then boosted with M306 G3 or M306 G3 KLH in incomplete Freund's adjuvant (IFA) . Two weeks after the boost the mice were bled for a 2° response. Two weeks after the 2° bleeding the mice were bled for a late 2° response. The mice were again boosted and bled 2 weeks later for a 3° response. Two weeks after the 3° bleeding, the mice were bled for a late 3° response. The boost schedule was repeated until the late 5° response was determined. All blood was collected in

Becton-Dickinson serum collection tubes and the serum separated out and used in direct and competition binding experiments.

The results of direct binding experiments using mouse immune serum are presented in Figures 3 and 4. Figure 3 represents an average value of assays performed after each bleed from 4 mice immunized with M306 G3. The figure legend lists the antigen which was bound to the plate. The control was bovine serum albumin (BSA). As can be seen clearly in Figure 3, immunization with M306 G3 mimic peptide led to the production of murine antibodies specifically reactive with both V3-loop and the M306 G3 peptide. The immunized mice had never been exposed to KLH and therefore the reactivity seen in Figure 3, middle bar, is due to specific reactivity with the mimic peptide. As can be seen in the later bleeds, particularly the 4° bleed, immunization with the mimic peptide results in the production of murine antibodies which are specifically reactive with the V3-loop peptide.

Figure 4 shows the results of a similar experiment in which M306 G3 conjugated to KLH was used to immunize 4 mice. The data presented in Figure 4 represents an average value for the 4 mice. The antigen which was fixed to the plate is shown in the legend. KLH was not present on the plate and therefore the middle bar represents reactivity toward the mimic peptide specifically. Again the antisera was reactive with V3-loop although a greater disparity was observed between the relative affinities of the antisera for V3-loop versus M306 G3 than was observed in the experiment described in the preceding paragraph.

Competition experiments were carried out using the mimic peptides and the mouse immune serum. As shown in Figure 5, in a first set of experiments, serum from a mouse immunized with M306 G3 KLH was mixed with peptides prior to incubation with V3-loop fixed to an ELISA plate. A significant decrease in the OD reading was observed when the serum was preincubated with either V3-loop or M306 G3 as compared with the OD reading when the serum was preincubated with PBS. This result indicates that mice immunized with the M306 G3 KLH conjugate produce antibodies which cross-react with both M306 G3 and V3-loop. Figure 6 shows the results of competition experiments in which M306 G3 BSA was fixed to the ELISA plate and the serum was from a mouse immunized with M306 G3 KLH. As can be seen in the figure, the M306 G3 peptide was able to compete effectively at antigen concentrations exceeding 10 μg/ml.

Competition experiments were also carried out using the affinity purified 28G12 monoclonal antibody. Figure 7 shows the results of a competition experiment with this antibody using ELISA plates to which V3-loop had been attached. Again, M306 G3 mimic peptide showed competitive behavior at concentrations exceeding 10 μg/ml. Figure 8 shows the results of a 28G12 competition experiment in which recombinant gρl20 was fixed to the ELISA plate. In this experiment, the inclusion of the M306 G3 peptide resulted in a substantial reduction in the OD determination indicating effective competition.

Equivalents

Those skilled in the art will know, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. These and all other equivalents are intended to be encompassed by the following claims.

Claims

1. A peptide having a primary amino acid sequence which is unrelated to the primary amino acid sequence of HIV gpl20, the peptide characterized by the ability to stimulate the production of anti-gpl20 antibodies when administered to a mammal.

2. A peptide of Claim 1 comprising the amino acid sequence:

AAl Gin AA2 Asn Arg AA3; wherein AAl, AA2 and AA3 represent any amino acid residue.

3. A peptide of Claim 2 wherein AAl and AA3 are selected from the group consisting of serine, threonine, methionine and cys eine and AA2 is selected from the group consisting of lysine, arginine, histidine, serine and threonine.

4. A peptide of Claim 3 comprising an amino acid sequence selected from the group consisting of: a) Ser Gin Thr Asn Arg Met; b) Met Gin Ser Asn Arg Ser; and c) Ser Gin Arg Asn Arg Ser.

5. A peptide of Claim 1 having a primary amino acid sequence which is unrelated to the primary amino acid sequence of HIV gpl20 V3-loop, the peptide characterized by the ability to stimulate the production of anti-V3-loop antibodies when administered to a mammal.

6. A peptide of Claim 4 comprising an amino acid sequence selected from the group consisting of:

Ser Gin Thr Asn Arg Met Ala; Met Gin Ser Asn Arg Ser

Ala; and

Ser Gin Arg Asn Arg Ser Ala, or fragments of (a), (b) or (c) .

7. A peptide of Claim 6 which is conjugated to an immunogenic carrier molecule.

8. A peptide of Claim 7 wherein the immunogenic carrier molecule is keyhole limpet hemacyanin.

9. A peptide conjugated to an immunogenic carrier molecule, the peptide having a primary amino acid sequence which is unrelated to the primary amino acid sequence of HIV gpl20, the conjugate characterized by the ability to stimulate the production of anti-gρl20 antibodies when administered to a mammal.

10. A peptide of Claim 9 comprising the amino acid sequence:

11. A peptide of Claim 10 wherein AAl and AA3 are selected from the group consisting of serine, threonine, methionine and cysteine and AA2 is selected from the group consisting of lysine, arginine, histidine, serine and threonine.

12. A peptide of Claim 11 comprising an amino acid sequence selected from the group consisting of: a) Ser Gin Thr Asn Arg Met; b) Met Gin Ser Asn Arg Ser; and c) Ser Gin Arg Asn Arg Ser.

13. A vaccine composition comprising a peptide in a physiologically acceptable carrier, the peptide having a primary amino acid sequence which is unrelated to the primary amino acid sequence of HIV gpl20 V3-loop, the peptide characterized by the ability to stimulate the production of anti-V3-loop antibodies when administered to a mammal.

14. A vaccine composition of Claim 13 further comprising an adjuvant.

15. A vaccine composition of Claim 13 wherein the peptide comprises an amino acid sequence selected from the group consisting of:

Asn Arg

Asn Arg or fragments of (a), (b) or (c) .

16. A vaccine composition of Claim 13 wherein the peptide comprises an amino acid sequence selected from the group consisting of: a) Ser Gin Thr Asn Arg Met; b) Met Gin Ser Asn Arg Ser; and c) Ser Gin Arg Asn Arg Ser.

17. A method for stimulating the production of an anti-HIV gpl20 immune response in a mammal comprising administering an effective amount of a vaccine composition comprising a peptide in a physiologically acceptable carrier, the peptide having a primary amino acid sequence which is unrelated to the primary amino acid sequence of HIV gpl20 V3-loop, the peptide characterized by the ability to stimulate the production of anti-V3-loop antibodies when administered to a^' mammal.

18. A method of Claim 17 wherein the mammal is a human.

19. A method of Claim 18 wherein the vaccine composition further comprises an adjuvant.

20. A method of Claim 17 wherein the peptide comprises an amino acid sequence selected from the group consisting of:

or fragments of (a), (b) or (c)

21. A method of Claim 17 wherein the peptide comprises an amino acid sequence selected from the group consisting of: a) Ser Gin Thr Asn Arg Met; b) Met Gin Ser Asn Arg Ser; and c) Ser Gin Arg Asn Arg Ser.

22. A method of Claim 20 wherein the peptide is conjugated to an immunogenic carrier molecule.

23. A method of Claim 22 wherein the immunogenic carrier molecule is keyhole linpet hemacyanin.

24. A peptide having: a) a primary amino acid sequence which is unrelated to the primary amino acid sequence of an essential protein of a virus; and b) the ability to interfere with a component involved in the function of the virus.

25. A peptide of Claim 24 wherein the essential protein of a virus is the HIV gpl20 protein.

26. A peptide of Claim 24 further characterized by the ability to stimulate the production of antibodies specifically reactive with the essential protein of a virus when administered to a mammal.

27. A peptide of Claim 26 wherein the essential protein of a virus is the HIV gpl20 protein.

28. A peptide of Claim 25 comprising the amino acid sequence:

29. A method for interfering with viral propagation comprising administering to an individual an effective amount of a therapeutic composition comprising a peptide in a physiological carrier, the peptide having: a) a primary amino acid sequence which is unrelated to the primary amino acid sequence of an essential protein of a virus; and b) the ability to interfere with a component involved in the propagation of the virus.

30. A method of Claim 29 wherein the essential protein of a virus is the HIV gρl20 viral protein.

31. A method of Claim 29 wherein the peptide is further characterized by the ability to stimulate the production of antibodies specifically reactive with the essential protein of a virus when administered to a mammal.

32. A method for screening an epitope library with a non-isolate specific antibody comprising the steps of: a) providing an epitope library containing bacterial cells infected with fusion phage; b) contacting the fusion phage with a biotinylated non-isolate specific antibody under conditions appropriate for binding of the non-isolate specific antibody to peptides displayed on the surface of the bacterial cells infected by the fusion phage; c) contacting the mixture from step b) with a streptavidin-coated substrate under conditions appropriate for the binding of biotin to streptavidin; d) removing non-specifically bound phage; e) eluting specifically bound phage; and f) infecting E_^. coli cells with the eluted phage.

33. A peptide conjugated to an immunogenic carrier molecule, the peptide having a primary amino acid sequence which is unrelated to the primary amino acid sequence of an essential protein of a virus, the conjugate having the ability to interfere with a component involved in the function of the virus and the ability to stimulate the production of antibodies specifically reactive with the essential protein of the virus when administered to a mammal.