Described herein are chimeric peptides comprising a soluble trimeric coiled-coil and all or a portion of the N-peptide region of HIV gp41. These molecules are stable, trimeric coiled-coils that inhibit HIV entry into cells, such as human cells. Such peptides can be further assessed to demonstrate their ability to serve as potent anti-HIV therapeutic molecules and thus, as therapeutic molecules or drugs.

SUMMARY OF THE INVENTION

Described herein are chimeric peptides comprising a soluble trimeric coiled-coil and all or a portion of the N-peptide region of HIV gp41. These molecules are stable, trimeric coiled-coils that inhibit HIV entry into cells, such as human cells. Such peptides can be further assessed to demonstrate their ability to serve as potent anti-HIV therapeutic molecules and thus, as therapeutic molecules or drugs.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to soluble peptides, referred to as soluble IQ (or IN) peptides, which, under the conditions described herein, fold into a stable trimeric coiled-coil (helical) structure and inhibit HIV infection of mammalian cells, such as human cells. In a specific embodiment, the soluble IQ peptides also bind D-peptide inhibitors of HIV infection of human cells, under the conditions described herein. In one embodiment, soluble IQ peptides comprise a trimeric coiled-coil peptide and a portion of the N-helix coiled-coil of HIV gp41 (HIV gp41 N-peptide); the components are present in the following "order": N-terminus--trimeric coiled-coil peptide--N-helix coiled coil of HIV gp41--C-terminus. The trimeric coiled-coil peptide can be from (comprise amino acid residues that correspond to those in) a variety of sources, such as GCN4, the yeast transcription activator; Moloney Murine Leukemia Virus (MoMLV); GCN4-pII, GCN4-pI.sub.QI and the ABC heterotrimer. It can also be from other designed trimeric coiled coils such as the isoleucine zipper (IZ) described by Tanaka et al., or derivatives of this `IZ` sequence. In those embodiments in which the trimeric coiled-coil peptide is from the isoleucine zipper, they are referred to as IZ peptides. Alternatively, it can comprise a trimeric coiled-coil peptide from HIV. Three examples of coiled-coils of interest are -- see Original Patent.

Tanaka's isoleucine zipper: Ac-YGGIEKKIEAIEKKIEAIEKKIEAIEKKIEA-NH.sub.2 (SEQ ID NO: 16) The "IZ" molecule derived from Tanaka et al., but with mutations: Ac-YGGIKKEIEQEAIEKIAIEKEIEA-NH2 (SEQ ID NO: 11).

The amino acid residues that comprise an IQ peptide of the present invention can be amino acid residues that are contiguous (consecutive) or noncontiguous (nonconsecutive) in the trimeric coiled-coil peptide from which it is derived and/or amino acid residues that are contiguous (consecutive) or noncontiguous (nonconsecutive) in HIV gp41 N-peptide, provided that the resulting IQ peptide (the IQ peptide in which they are present) is stable, soluble, helical, and trimeric and inhibits HIV infection of human cells. In the embodiments of IQ peptides in which nonconsecutive amino acid residues of either or both components of the IQ peptide are present, the residues, as included in the IQ peptide, can be consecutive or can be separated or joined by a linker. The linker can be, for example, an amino acid residue(s) that do not occur between two amino acid residues in the peptide from which the component is derived. Alternatively, the "linker" can be a chemical or synthetic linker. A component of an IQ peptide of the present invention is considered to be "derived from" another peptide (e.g., a trimeric coiled-coil or HIV gp41 N-peptide) if the component itself (or the nucleic acid molecule(s) that encode the amino acid sequence) is obtained or isolated/separated from a source in which it occurs (e.g., from a cell in which the peptide occurs, such as a portion of a protein from which it can be removed) or is produced by recombinant DNA methods, chemical synthesis or any other method, to comprise an amino acid sequence or a nucleic acid sequence that is the same as or substantially the same as the sequences of the peptide. That is, the term is intended to be interpreted broadly and does not require that a component be physically derived from the peptide referred to.

In the embodiments in which the soluble IQ peptides comprise an IQ region that is a GCN4 trimeric coiled-coil peptide, they are referred to as IQN peptides. IQN peptides comprise all or a portion of GCN4-pI.sub.Q I (formerly referred to as GCN4-pIQ in U.S. Provisional Application No. 60/101,058; Eckert D. M. et al. J. Mol. Biol, 284: 859 865 (1998)) or a modified version of all or a portion of GCN4-pI.sub.QI, such as a modified portion that includes mutations for increased solubility, and all or a portion of the HIV gp41 N-peptide. Typically, 5 or more (e.g., 7, 8, 9 or 10) amino acid residues from HIV-gp41 N-peptide up to and including all of the residues of the N-peptide will comprise the HIV gp41 component of the IQ peptides.

Soluble IQN peptides of the present invention comprise, in specific embodiments, a portion of the HIV gp41 N-peptide sufficient to bind the C-peptide (region) of HIV gp41 and a sufficient portion of the GCN4 trimeric coiled-coil peptide or a modified version of the GCN4 peptide that the resulting IQN peptide is a soluble trimeric (helical) coiled coil. In further embodiments, IQN peptides comprise a portion of the HIV gp41 N-peptide that includes the amino acid residues which form the pocket or cavity of HIV gp41 (the pocket-comprising residues of the N-peptide). In yet further embodiments, IQN peptides do not comprise amino acid residues which form the pocket or cavity of HIV gp41. They do, however, comprise amino acid residues from HIV gp41. (See, for example, IQN23, IQN36 and IQN26).

Nomenclature of IQ peptides refers to the number of amino acid residues from the HIV gp41 N-peptide or a modified version of HIV gp41 N-peptide present in the IQ peptide. For example, 17 amino acid residues of HIV gp41 are included in the IQN17 peptide described herein. As explained above, the trimeric coiled coil peptide component of an IQ peptide must be sufficient in amino acid composition (identity and number/length) to result, when joined to the HIV gp41 N-peptide portion, in formation of a soluble trimeric helical (coiled-coil) IQ peptide. In certain embodiments of the IQN peptides of the present invention, the trimeric coiled-coil peptide, referred to as the "GCN4 portion", comprises at least 15, 16, 17, 18, 19 or 20 amino acid residues of GCN4. The amino acid residues present in the components of an IQN peptide can correspond to amino acid residues that are contiguous (consecutive) or noncontiguous (nonconsecutive) in, respectively, the GCN4 transcription activator (or GCN4-pI.sub.QI) and HIV gp41 N-peptide or a modified version of the activator or the N-peptide, provided that the resulting IQN peptide is an inhibitor of HIV infection of human cells, as described herein. The IQ and IZ peptides of the present invention can be produced as a continuous peptide or as components that are joined or linked after they are formed. As used herein, the terms "joined" or "joined in such a manner" or "incorporated" include incorporating amino acid residues by either approach.

For example, the GCN component of an IQN peptide can comprise consecutive amino acid residues from GCN4-pI.sub.QI, modified, if desired (e.g., to increase solubility, as is the case in IQN17 (SEQ ID NO.: 1)). Alternatively, amino acid residues that are not consecutive in the GCN4 activator (or in GCN4-pI.sub.QI), joined in such a manner that they are nonconsecutive or consecutive in the resulting GCN4 component of an IQN peptide, can be incorporated in the IQN peptide. Similarly, the amino acid residues of the HIV gp41 N-peptide component of an IQN peptide of the present invention can be amino acid residues that occur consecutively or nonconsecutively in HIV gp41 N-peptide and can be incorporated into in IQN peptide in such a manner that they are consecutive or nonconsecutive in the resulting peptide. In the embodiments in which nonconsecutive amino acid residues are used, they can be separated by one or more "linker" molecules, if needed to retain the respective functions/characteristics of the components and of the IQN peptide. For example, an amino acid residue(s) other than the residue(s) that normally occur between two amino acid residues of GCN4 or HIV gp41 N-peptide can be used to link or join the two amino acid residues in the IQN peptide. Alternatively, the linker can be a chemical or synthetic linker, for example. Under the conditions described herein, IQN peptides have been shown to fold into a stable structure, bind peptide inhibitors of HIV-1 infection and inhibit HIV infection of human cells. For example, IQN17 and IQN23 have been shown to fold into stable structures, bind D-peptides previously shown to be inhibitors of HIV-1 infection and inhibit HIV infection of human cells. IQN36, as well as versions of IQN17 that are shortened in the `IQ` region are also described. These shortened versions may be therapeutically advantageous because, for example, they are easier and less expensive to produce than are larger peptides.

A specific embodiment of an IQN peptide is IQN17, which contains 29 residues of GCN4-pI.sub.QI, including three mutations for increased solubility, and 17 residues of HIV; there is a one residue overlap between the two proteins, making the total length of the fusion protein 45 residues. The sequence of GCN4-pIqI is ac-MKQIEDKIEEILSKQYHIENEIARIKKLIGER (SEQ ID NO: 17). In this embodiment, the HIV Sequence is: LLQLTVWG IKQLQARIL (SEQ ID NO: 18). The sequence of IQN17 is: ac-RMKQIEDKIEEIESKQKKIENEIARIKK LLQLTVWGIKQLQARIL-am (SEQ ID NO.: 1). In the sequences presented, ac represents an N-terminal acetyl group and am represents a C-terminal amide group. IQN17 has been shown to inhibit HIV of human cells, as described herein.

Shortened versions of IQN17, which each contain 17 amino acid residues of HIV gp41 N-helix (SEQ ID NO.: 2), but include a shorter GCN component than is present in IQN17, are also the subject of this invention. Specific examples of these shortened IQN17 peptides are: a) shortened IQN17 #1, (SEQ ID NO.: 5), in which there are eight amino acid residues of GCN4-pI.sub.QI: EIARIKKL (SEQ ID NO.: 19); b) shortened IQN17 #2 (SEQ ID NO.: 6), in which there are 15 amino acid residues of GCN4-pI.sub.QI: KQKKIENEIAAIKKL (SEQ ID NO.: 20) and c) shortened IQN17 #3 (SEQ ID NO.: 7), in which there are 15 non-HIV amino acid residues KIKKIENEIARIKKL (SEQ ID NO.: 21). This is GCN4-pI.sub.QI' with an I to Q mutation, and is referred to as GCN4pII'. d) shortened IQN17 #4 (SEQ ID NO.: 8), in which there are 21 amino acid residues of GCN4-pI.sub.Qp: KIEEIESKQKKIENEIARIKKL (SEQ ID NO.: 22 and e) shortened IQN17 #5 (SEQ ID NO.: 9), in which there are 21 non-HIV amino acid residues: KIEEIESKIKKIENEIARIKK (SEQ ID NO.: 23).

Another specific embodiment of this invention is IQN23. One embodiment, referred to as IQN23 version 1, has the following sequence: ac-RMKQIEDKIEEILSKQYHIENEIARIKKLIEAQQHLLQLTVWGIKQLQARIL-am (SEQ ID NO.: 2). In IQN23 version 1, there are 29 amino acid residues in the GCN4 component and 23 amino acid residues in the HIV gp41 component (a total of 52 amino acid residues). A second embodiment of an IQN23, referred to as IQN23 version 2, also includes 23 amino acid residues of N-peptide of gp41 and 29 amino acid residues in its GCN component, but differs from IQN23, version 1 at amino acid residues 17 and 18 (and the L at position 15 is changed to E). In version 1, these two residues are, respectively, Y and H and in version 2, they are, respectively, both K. Three modifications have been made--the L, Y and H. These are the same three modifications that were made to make IQN17 more soluble. The two "versions" are referred to as GCN4-pI.sub.QI and GCN4-pI.sub.QI'. The sequence of IQN23 version 2 is: ac-RMKQIEDKIEEIESKQKKIENEIARIEAQQHLLQLTVWGIKQLQARILNH2 SEQ ID NO.: 3). As described herein, IQN23 (both versions) inhibits HIV infection more effectively than does IQN17.

Another specific embodiment of IQN peptides of this invention is IQN36, in which there are 30 amino acid residues of GCN4-pI.sub.QI and 36 amino acid residues of HIV. The sequence of IQN36 (SEQ ID NO.: 4) is shown in FIG. 2. The sequence of the GCN4-pI.sub.QI component is: ac-RMKQIEDKIEEIESKQKKIENEIARIKKLI (SEQ ID NO.: 24) and the HIV amino acid residues are: SGIVQQQNNLLRAIEAQQHLLQLTVWGIKQLQARIL--NH2 (SEQ ID NO.: 25). The sequence of IQN36 is: ac-RMKQIEDKIEEIESKQKKIENEIARIKKLI SGIVQQQNNLLRAIEAQQHLLQLTVWGIKQLQARIL--NH2 (SEQ ID NO.: 3).

A wide variety of fusion proteins which are variants of IQN17 can be produced and used to inhibit HIV. Fusion proteins include proteins made as a single continuous molecule or as components that are subsequently joined or linked together. Any of a wide variety of variations can be made in the GCN4-pIqI component of IQN17 and used in the method, provided that these changes do not alter the trimeric state of the coiled-coil. For example, the amino acid composition of the GCN4 component can be changed by the addition, substitution, modification and/or deletion of one or more amino acid residues, provided that the trimeric state of the coiled-coil is maintained. For example, the Asp residue in IQN17 (at an "f-position" of the coiled coil) can be replaced by any of the naturally-occurring amino acids. (O'Neil and DeGrado, Science 250:646 (1990)). Alternatively, this component of the fusion protein can be a trimenic version of the coiled-coil region of another protein, such as that from Moloney Murine Leukemia Virus (Fass, D. et al. Nature Struct. Biology, 3:465 (1996)), GCN4-pII (Harbury et al., Nature, 317:80, 1994) or the ABC heterotrimer (Nautiyal and Alber, Protein Science 8:84 (1999)), or the isoleucine zipper described by Tanaka, et al.

Changes can also be made in the amino acid composition of the fusion protein component which is the C-terminal portion of the HIV gp41 N peptide to produce a variety of fusion proteins to be used to prevent HIV infection of cells. The C-terminal portion can be changed by the addition, substitution, modification and/or deletion of one or more amino acid residues. The amino acid composition of either or both components of the fusion protein can be altered, and there is no limit to the number or types of amino acid residue changes possible, provided that the trimeric state of the coiled-coil is maintained. It is not necessary that the pocket or cavity of gp41 be included, although in many embodiments the pocket is present.

In all embodiments, controlled or time release (gradual release, release at a particular time after administration or insertion) of the drug can be effected by, for example, incorporating the drug into a composition which releases the drug gradually or after a defined period of time. Alternatively, the drug can be incorporated into a composition which releases the drug immediately or soon after its administration or application (e.g., into the blood, vagina, mouth or rectum). Combined release (e.g., release of some of the drug immediately or soon after insertion, and over time or at a particular time after insertion) can also be effective (e.g., by producing a composition which is comprised of two or more materials: one from which release or delivery occurs immediately or soon after insertion and/or one from which release or delivery is gradual and/or one from which release occurs after a specified period). For example, a drug or drugs which bind the HIV cavity can be incorporated into a sustained release composition such as that taught in U.S. Pat. No. 4,707,362. The cream, foam, gel or suppository can be one also used for birth control purposes (e.g., containing a spermicide or other contraceptive agent), although that is not necessary (e.a., it can be used solely to deliver the anti-HIV drug, alone or in combination with another non-contraceptive agent, such as an antibacterial or antifungal drug or a lubricating agent). An anti-HIV drug of the present invention can also be administered to an individual through the use of a contraceptive device (e.g., condom, cervical cap, diaphragm) which is coated with or has incorporated therein in a manner which permits release under conditions of use a drug or drugs which bind the HIV gp41 N-helix coiled coil. Release of the drug(s) can occur immediately, gradually or at a specified time, as described above. As a result, they make contact with and bind HIV and reduce or prevent viral entry into cells.

Fusion proteins of the present invention comprise a soluble, trimeric form or version of a coiled-coil, such as a soluble, trimeric form or version of a coiled-coil region of a protein (of non-HIV origin or of HIV origin) and a sufficient portion of the C-terminal end of the N-peptide of HIV gp41 to bind to the C-peptide region. In one embodiment, the portion of the C-terminal end of the N-peptide comprises sufficient amino acid residues to bind to the C-peptide region and include the HIV coiled-coil cavity or hydrophobic pocket (the pocket comprising residues of the N-peptide). The N-peptide of HIV gp41 can be that of HIV-1, HIV-2, another HIV strain or a strain from another species (e.g., simian immunodeficiency virus (SUV), feline immunodeficiency virus or Visna virus). For example, HIV-2 sequence LLRLTVWGTKNLQARVT (SEQ ID NO: 26), SIV sequence LLRLTVWGTKNLQTRVT (SEQ ID NO: 27) or a sequence comprising invariant residues in HIV-1, HIV-2 and SIV (represented LLXLTVWGXKXLQXRXX (SEQ ID NO: 28), wherein amino acid residues L, T, V, W, G, K, Q, and R are the single letter code used for amino acid residues and X can be any amino acid residue). Also the subject of this invention is a soluble trimeric model of the HIV gp41 hydrophobic pocket, which can be a D-peptide or an L-peptide and comprises a soluble trimeric coiled-coil and a sufficient portion of the N-peptide region of HIV gp41 to comprise the amino acid residues which form the pocket of the N-helix coiled-coil region of HIV gp41. The D- or L-peptide can comprise, as the soluble, trimeric coiled-coil, the coiled-coil of GCN4-pI.sub.QI; GCN4-pII; Moloney Murine Leukemia Virus or the ABC heterotrimer. The component which is a sufficient portion of the N-peptide of HIV gp41 to comprise the amino acid residues of the pocket can comprise, for example: LLQLTVWGIKQLQARIL of HIV-1(SEQ ID NO: 18); LLRLTVWGTKNLQARVT of HIV-2 (SEQ ID NO: 26); LRLTVWGTKNTLQTRVT of SIV (SEQ ID NO: 27) or the invariant residues of these, which are: LLXLTVWGXKXLQXRXX (SEQ ID NO: 28).

One embodiment of the instant invention is fusion proteins in which the components are a trimeric version of the coiled-coil region of a protein (such as GCN4-pI.sub.QI) and the N-helix coiled-coil of HIV gp41 that include all, part or none of the N-helix cavity. That is, a fusion protein of the present invention can comprise a trimeric form of the coiled-coil region of GCN4-pI.sub.QI and a portion of the N-peptide of HIV-1 gp41, wherein the portion of the N-peptide of gp41 comprises part, or all, or none of the N-helix cavity of HIV-I gp41. For example, a fusion protein can be made that contains residues from GCNL4-pI.sub.QI and residues from N36. The fusion protein, denoted IQN24n, contains 29 residues of GCN4-pI.sub.QI, including three mutations for increased solubility, and 24 residues from the N-terminal end of N36 (SGIVQQQNNLLRAIEAQQHLLQLT) (SEQ ID NO: 29); for recombinant expression in E. coli, an extra Met residue is included at the N-terminus. For example, a fusion protein can comprise a portion of the N-peptide of HIV gp41 comprising the amino acid sequence of (SEQ ID NO: 29). The sequence of IQN24n is: MRMKQIEDKIEEIESKQKKIENEIARIKKLISGIVQQQNNLLRAIEAQQHLLQLT (SEQ ID NO: 30). This fusion protein can be made by a variety of methods, including chemical synthesis or recombinant DNA methods or by recombinant expression in E. coli, in which case the N- and C-termini are not blocked. Because the superhelix parameters of the GCN4-pIQI coiled-coil are nearly identical to the HIV gp41 N-helix coiled-coil, the resulting fusion protein molecule IQN24n is predicted to form a long trimeric coiled-coil, which presents part of the gp41 N-helix coiled-coil as a trimer (not aggregated).

The same strategy described herein to solve this problem for the gp41 hydrophobic pocket can be applied towards the development of soluble, trimenic models of the gp41 N-helix coiled-coil region in general. Such trimeric models (including IQN 17, but also including, for example, peptides that do not contain the pocket residues of gp41) can be used as inhibitors.

Any of a wide variety of variations can be made in the GCN4-pIQI component of fusion proteins described herein (e.g., IQN17 or IQN24n) and used in the method, provided that these changes do not alter the trimeric state of the coiled-coil. Changes can also be made in the amino acid composition of the fusion protein component which is the portion from the HIV gp41 N36 peptide, to produce variants (e.g., variants of IQN17 or IQN24n). There is no limit to the number or types of amino acid residue changes possible, provided that the trimeric state of the coiled-coil and the structure of the surface of the fusion protein corresponding to the N-peptide coiled coil of HIVgp41 are maintained. The fusion protein component which is the portion of the HIV gp41 peptide can include all, part, or none of the N-helix cavity. For example, other parts of N51, N36, DP-107, or other regions of the HIV gp41 N-helix region can be fused to GCN4-pI.sub.QI (or another trimeric version of the coiled-coil region of a protein) to generate trimeric (not aggregated) helical coiled-coil fusion proteins and used in the method. There is no limit to the number or types of fusion proteins that can be designed and generated, provided that the trimeric state of the coiled-coil and the structure of the surface of the fusion protein corresponding to the N-peptide coiled coil of HIV gp41 are maintained. Such fusion proteins can be designed and generated using methods known to those of skill in the art, such as evaluating heptad-repeat positions or superhelix parameters of coiled coils.

IQN17 is useful as an anti-HIV therapeutic agent, a prophylactic agent or drug to prevent HIV infection, a reagent for identification (screening for) or designing other anti-HIV therapeutics or prophylactics, and an immunogen to elicit antibodies that prevent HIV infection.

Applicants have shown that a portion of the N-peptide can be solublized by addition of a soluble, trimeric coiled-coil, GCN4-pI.sub.QI. The resulting molecule is stable under physiological conditions and is correctly folded such that IQN17 presents a surface that is structurally complementary to the C-peptide region of HIV gp41. Further, IQN17 and similar molecules can be assessed for their ability to bind to the C-helical region of gp41, and inhibit its function. The N-helical core of gp41 is highly conserved (in terms of amino acid composition) and thus, it is likely that IQN17 and variants thereof will be broadly neutralizing against a variety of clinical HIV strains and, thus, useful therapeutically.

IQN17, which is based upon the known structure of the gp41 ectodomain, consists, in one embodiment, of three N-peptides joined to (or present in larger molecule with) a soluble trimeric coiled-coil and arranged to fold into a substantial part of the N-helical core with peptide biding sites of the N-peptides exposed.

IQN17 protein can be produced by a variety of methods. For example, it can be chemically synthesized. Alternatively, it can be produced, using known methods and expression systems, by expressing IQN17 protein-encoding DNA, which can be a single DNA that encodes the entire IQN17 protein. Alternatively, protein synthetic methods can be used to produce IQN17 protein.

IQ(IN) peptides can have a wide variety of sequences, both in the N-helix and fused coiled-coil components, and can be comprised of L-amino acid residues, D-amino acid residues and modified amino acids residues. IQN17 can include amino acid residues in addition to those of the helices and the fused coiled-core (e.g., to stabilize the molecule). It is likely that the IQN17 described here can be altered to enhance stability and activity. Minor changes in the fused coiled-coil and the exact borders of the N-Helix are likely to have significant effects on the stability, yield, and activity of IQN17.

As currently constructed, IQN17 exposes a portions of three C-peptide binding sites. A strategy for exposing longer segments of the C-peptide binding site on IQN17 (or related molecules) involves extending the N-peptide region of IQN17.

IQN17 is useful in a variety of contexts. As described herein, IQN17 is a potent inhibitor of viral membrane fusion, and, thus, acts on the virus before it enters the cell (unlike current practical therapy, which acts on HIV-infected cells). IQN17 is quite soluble and has been shown to be stable under the conditions described herein. It is reasonable to expect that its size will prevent rapid filtration in the kidney. In addition, IQN17 dimers can be made by disulfide crosslinking, to produce a molecule filtered to a lesser extent than the IQN17 "monomer". Thus, it is reasonable to expect that dimers have an enhanced bioavailability when compared to the C-peptides.

IQN17 prevents virus from entering cells, unlike standard therapy that targets viral proteins after viral entry, and thus, IQN17 can be used prophylactically to prevent infection or reduce the extent to which infection occurs. One use for such a therapeutic is in the event of a needlestick injury, such as might occur in a hospital or in settings in which needles contaminated with HIV are shared.

In one embodiment of the present invention, IQN17 is used to reduce HIV infection in an individual. In this embodiment, IQN17 is administered, either as IQN17 itself or via expression of IQN17-encoding DNA in appropriate host cells or vectors, to an individual in sufficient quantity to reduce (totally or partially) HIV infection of the individual's cells. That is, a dose of IQN17 sufficient to reduce HIV infection (an effective dose) is administered in such a manner (e.g., by injection, topical administration, intravenous route that it inhibits (totally or partially) HIV entry into cells. In one embodiment, a gene therapy approach is used to provide the effective dose, by introducing cells that express IQN17 protein into an individual. IQN17 can be administered to an individual who is HIV infected, to reduce further infection, or to an uninfected individual, to reduce infection.

The serum stability of IQN17 can be tested, using known methods to ascertain its therapeutic potential.

The outside surface of the fusion coiled-coil of IQN17 can be varied, for example, to enhance bioavailability, decrease toxicity, and avoid immune clearance. IQN17 exhibits potent inhibitory activity and GCN4-pI.sub.QI does not, it is the exposed N-peptide region that is responsible for inhibition. The rest of the molecule provides a scaffold for displaying the N-peptide. Therefore, this scaffold can be modified without adversely affecting the inhibitory activity of IQN17. Modification of the scaffold may provide several advantages. First, it would facilitate procedures in which multiple administrations of IQN17 are required. For example, when IQN17 is used as an anti-HIV therapeutic agent, multiple doses might be required. After extended administration, individuals might develop antibodies to IQN17 which are likely to increase its clearance from the body. The availability of multiple versions of IQN17 would help to circumvent this problem by evading preexisting antibodies. Second, it may be possible to design versions of IQN17, for example by introducing glycosylation sites on the external surface, in which the scaffold is less immunogenic.

The trimer of helical hairpins (TOH) is a common feature of many viral membrane fusion proteins (Singh, M. et al. J. Mol. Biol.290,1031 1041(1999)). It has been observed in crystal structures of influenza, Ebola SV5 (simian parainfluenza virus 5), and RSV (human respiratory syncitial virus). In addition, many other members of the retrovirus, paramyxovirus, and filovirus families are predicted to contain this motif. A similar structure has been observed in the associated vertebrate vesicle fusion proteins and may be found in sperm-egg, fertilization proteins. The basic strategy described herein can be applied to any of these systems in order to inhibit fusion.

 

Claim 1 of 2 Claims

1. A soluble trimeric coiled-coil peptide comprising a peptide IZN26, wherein the amino acid sequence of IZN26 is: TABLE-US-00003 (SEQ ID NO:14) -- see Original Patent.

 

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