Inhibition of binding of complement Factor H

Details Inhibition of binding of complement Factor H Inhibition of binding of complement Factor H

1996-12-30

2001-11-20

Celsa, Bennett (Department: 1627)

Drug, bio-affecting and body treating compositions

Designated organic active ingredient containing

Peptide containing doai

C514S009100, C514S010100, C514S011400, C514S012200, C514S013800, C514S014800, C514S017400, C530S317000, C530S324000, C530S325000, C530S326000, C530S327000, C530S328000, C530S329000, C530S330000, C530S350000

Reexamination Certificate

active

06319890

ABSTRACT:

BACKGROUND OF THE INVENTION
(1) Field of the Invention
This invention relates generally to peptide chains used to block binding sites on pathogens and more particularly to peptide chains used to block binding sites for Complement Factor H (CFH) on pathogens such as the Human Immunodeficiency Virus (HIV).
(2) Description of Related Art
Various pathogens, like HIV, have evolved the ability to escape from complement-mediated lysis. See, e.g., Marschang P. Sodroski J., Wurzner R. and Dierich M. P., “Decay-accelerating Factor (CD55) Protects Human Immunodeficiency Virus Type 1 From Inactivation by Human Complement,” Eur. J. Immunol. 25: 285-90 (1995) and Horstmann, R. D., Sievertsen, H. J., Knobloch, J. and Fischetti, V. A., “Antiphagocytic Activity of Streptococcal M Protein: Selective Binding of Complement Control Protein Factor H,” Proc. Nat'l. Acad. Sci. U.S.A. 85: 1657-1661 (1988). The complement system is part of the immune system. Its main functions are the opsonisation of micro-organisms to allow more efficient phagocytosis and complement-mediated lysis of pathogens. To turn down the activity of complement and to protect host cells from unspecific damage, regulators of complement activity (RCA's) are required. Among this family of negative regulators is CFH, a very abundant plasma protein. CFH has been shown to down-regulate the alternative pathway of complement activation by directly affecting the formation and stability of C3 and C5 convertase and by acting as a cofactor for the cleavage of C3b into its inactive form iC3b as described by Whaley, K. and Ruddy, S., “Modulation of the Alternative Complement Pathway by &bgr;1H Globulin,” J. Exp. Med. 144: 1147 (1976) and Pangburn, M. K., Schreiber and Eberland, H. J. Muller, “Human Complement C3b Inactivator: Isolation, Characterization and Demonstration of an Absolute Requirement for the Serum Protein &bgr;1H for Cleavage of C3b and C4b in Solution,” J. Exp. Med. 146:257 (1977). Responsible for these effects is a functional site located in the first 5 short consensus repeats (SCR 1-5) of CFH as described in Alsenz, J., Lambris, J. D., Schulz, T. F. and Dierich, M. P., “Localization of the Complement Component C3b-binding Site and the Cofactor Activity for Factor I in the 38-kDa Tryptic Fragment of Factor H,” Biochem. J. 224: 389 (1984) and Gordon, D. L., Kaufman, R. M., Blackmore, T. K., Kwong, J. and Lublin, D. M., “Identification of Complement Regulatory Domains in Human Factor H,” J. Immunol. 155: 348 (1995). An interaction site for polyanionic molecules has been mapped in SCR 13. Pangburn, M. K., Atkinson, M. A. and Meri, S., “Localization of the Heparin-binding Site on Complement Factor H,” J. Biol. Chem. 266: 16847 (1991). For example, SCR 13 has been shown to bind streptococcal protein M and is responsible for protection of M+ strains from complement-mediated bactericidal activity. See Horstmann et al. article, above. Recently, additional binding regions for negatively-charged particles in SCR 7 have also been described. Blackmore, T. and Gordon, D., “SCR 7 is a Major Heparin/Sialic Acid Binding Site of Complement Factor H,” J. Mol. Immunol. 33 (S1): 15 (1996). It is believed that SCR 7 is a potential binding site for host cells.
Applicants have recently shown the CFH interacts with specific sites on the envelope of HIV. Specifically, envelope glycoproteins, gp120 and gp41, have been demonstrated to be binding sites for CFH. See Stoiber, H., Ebenbichler, C., Schneider, R., Janatova, J. and Dierich, M. P., “Interaction of Several Complement Proteins with gp120 and gp41, the Two Envelope Glycoproteins of HIV-1,” AIDS 9:19 (1995) and Pinter, C., Siccardi, A. G., Longhi, R. and Clivio, A., “Direct Interaction of Complement Factor H with the C1 Domain of HIV Type 1 Glycoprotein 120,” AIDS Research & Human Retroviruses 11: 577 (1995). Incubation of free HIV or HIV-infected cells with sera that has been depleted of CFH (NHS
CFH—
) results in the complement-mediated virolysis of primary HIV isolates and the killing of HIV-infected cells. Re-titration of CFH to NHS
CFH—
reconstitutes the resistance of HIV against complement-mediated destruction. See Stoiber, H., Pinter, C., Siccardi, A. G., Clivio, A. and Dierich, M. P., “Efficient Destruction of Human Immunodeficiency Virus in Human Serum by Inhibiting the Protective Action of Complement Factor H and Decay Accelerating Factor (DAF, CD55),” J. Exp. Med. 183: 307 (1996).
In addition, Applicants have shown that the antibody 2F5, described by Katinger et al. (See W095/07354 A1) interacts with a CFH binding site in gp41. From this information, the Applicants hypothesized that one of the HIV-neutralizing effects of this antibody is due to the inhibition of CFH interaction with gp41, resulting in complement-mediated lysis of the virus.
The present invention is based on the assumption that if the hypothesis described above is correct, it is not necessary to use an antibody for specifically blocking the binding sites on pathogens. Using a molecule which is able to block sites on pathogens which act as binding sites for CFH should have the same or even more promising effects.
SUMMARY OF THE INVENTION
It is accordingly an object of the invention to eliminate the need for antibody-mediated pathogen destruction. It is a further object of the invention to provide a pathogen specific peptide chain that binds to the sites that would otherwise be available to bind CFH so that CFH-mediated lysis of pathogens is not prevented.
It is a still further object of the invention to provide a peptide chain that does not have the active sites of CFH and does not have the binding sites on CFH that bind to host cells to regulate complement activity.
By screening with specific peptides, Applicants have discovered that CFH binding on pathogens can be inhibited so that complement-mediated destruction of the pathogens is induced. The Applicants have discovered that binding inhibition can be accomplished with peptides containing less than 100 amino acids. The amino acid chains, SEQ. ID NO: 1, were derived from the SCR 13 region of CFH. Whole CFH has been modified so that the areas involved in mediating the interaction with pathogens are conserved while the areas responsible for inhibition of complement-mediated lysis are deleted. Additionally, areas believed to be involved in binding to host cells such as SCR7 have also been deleted to provide a pathogen-specific peptide chain.
Such pathogen-specific peptide chains can be synthesized in a variety of ways including mutation and expression of mutated CFH DNA. Using known peptide synthesis procedures, it is even possible to use cyclic peptides or introduce D-amino acids which can result in higher resistance of the peptide chain to degradation by human serum.
By screening peptide and pseudopeptide libraries, the invention offers the possibility to detect in addition to the described peptides, other molecules which interfere with CFH binding on various pathogens. The effect of those substances can be easily checked by incubating pathogen which are resistant to human serum with antibodies specific against the pathogen, serum and peptides or pseudopeptides followed by testing and analyzing for the presence of complement-mediated lysis of the pathogen.


REFERENCES:
patent: 5472939 (1995-12-01), Fearon et al.
patent: 6074645 (2000-06-01), Diamond et al.
patent: WO 95/07354 (1995-03-01), None
Lellouch et al., Biochemiistry vol. 31 pp. 2279-2285, 1992.*
Ripoche et al., “The Complete Amino Acid Sequence of Human Complement Factor H”, Biochem J., (1988), v. 593, pp. 593-602.*
Journal of Biological Chemistry, vol. 266, No. 25, Sep. 5, 1991, MD US, pp. 16847-16853, MK Pangburn et al.: “Localization of heparin-binding site on complement factor H”.
Proccedings of the National Academy of Sciences of USA, vol. 93, No. 20, Oct. 1, 1996, Washington US pp. 10996-11001, AK Sharma & MK Pangburn: “Identification of three physically and functionally distinct binding sites for C3b in human complement factor H by deletion mutagenesis”.
AIDS Research and Human Retrovirus

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