Patent EA-002688-B1 (EA)
Title: Method for inactivating target cells agents and compositions causing cytolysis and compounds used for producing said agents
Inventor: SOEGAARD MORTEN
IPC: A61K47/48
Filed: 1998-07-02
Abstract: 1. A method for inactivating target cells in the presence of T cells by bringing the two types of cells in contact with a superantigen (SAG) in the presence of an immune modulator, characterized in that at least one of the superantigen and the immune modulator is conjugate with targeting moiety and targeting moiety is selected from the group, consisting of antibody, an antigen binding fragment thereof, Fab-fragment antibody or Fab2-fragment antibody or a single chain antibody, and said superantigen is modified to have a decreased ability to bind to MRC class II antigen and decreased seroreactivity and immunogenicity in human blood serum compared to the corresponding wild type superantigen; and said immune modulator is selected from the group consisting of cytokines, chemokines and extracellular parts of lymphocyte surface bound receptors and ligands. 2. The method of Claim 1, characterized in that the superantigen and the immune modulator are both conjugated with the same targeting moiety in a form of a triple conjugate. 3. The method of Claim 1, characterized in that only the superantigen and targeting moiety are conjugated, whereas the immune modulator is not conjugated. 4. The method of Claim 1, characterized in that the target cells are inactivated in vivo in an individual suffering from a disease associated with the target cells. 5. The method of Claim 4, characterized in that the disease is a cancer. 6. The method of Claim 1, characterized in that the superantigen is a chimera, comprising succession, derivative from two or more wild-type superantigens. 7. The method of Claim 1, characterized in that the immune modulator is IL-2. 8. The method of Claim 1, characterized in that the immune modulator is an extracellular part of a B7 molecule. 9. The method of Claim 1, characterized in that an extracellular part of a B7 molecule is selected from the group, comprising CD80 and CD86. 10. The method of Claim 1, characterized in that that the immune modulator has been modified to show a decreased affinity for its lymphocyte receptors, compared to the corresponding native form. 11. A superantigen conjugate complying with the formula (T)x(Sag)y(IM)z wherein y>0 and z>0; T is a targeting moiety, selected from the group, consisting of antibody, an antigen binding fragment thereof, Fab-fragment antibody or Fab2-fragment antibody or a single chain antibody, Sag corresponds to superantigen, which has been modified in such a way as to have a decreased ability to bind to MRC class II antigen and decreased seroreactivity and immunogenicity in human blood serum compared to the corresponding wild type superantigen; IM is immune modulator which is selected from the group consisting of cytokines, chemokines and extracellular parts of lymphocyte surface bound receptors and ligands. 12. A superantigen conjugate of Claim 11, characterized in that 10>x>0, 10>y>1 and 10>z>1. 13. A superantigen conjugate of Claim 12, characterized in that x, y and z are integers 1-3. 14. A superantigen conjugate of Claim 11, characterized in that T comprises at least T' and T'', superantigen is fused C-terminally to T' and the immune modulator is fused C-terminally to T''. 15. A superantigen conjugate of Claim 11, characterized in that the superantigen is a chimera, comprising succession, derivative from two or more wild-type superantigens. 16. A superantigen conjugate of Claim 11, characterized in that the immune modulator is IL-6. 17. A superantigen conjugate of Claim 11, characterized in that the immune modulator is an extracellular part of a B7 molecule. 18. A superantigen conjugate of Claim 17, characterized in that an extracellular part of a B7 molecule is selected from the group, comprising CD80 and CD86. 19. A superantigen conjugate of Claim 11, characterized in that that the immune modulator has been modified to show a decreased affinity for its lymphocyte receptors, compared to the corresponding native form. 20. A superantigen conjugate of Claim 14, wherein superantigen is Staphylococcal enterotoxin A, T' is the CH1-domain of C215 Fab fragment, T'' is the light chain of the C215 antibody and immune modulator is IL-2. 21. A superantigen conjugate of Claim 14, wherein superantigen is fused to T' via a flexible hydrophilic amino acid linker B of 3-11 amino acid residues and immune modulator is fused to T'' via a hydrophilic and neutral or positively charged amino acid linker Q of 10-20 amino acid residues. 22. A superantigen conjugate of Claim 21, wherein B is selected from the group consisting of Gly-Gly-Pro and Pro-Ala-Ser-Gly-Gly-Gly-Gly-Ala-Gly-Gly-Pro (SEQ ID N 19), and Q is selected from the group consisting of Gly-Pro-Arg-Gln-Ala-Asn-Glu-Leu-Pro-Gly-Ala-Pro-Ser-Gln-Glu-Glu-Arg (SEQ ID N 23), Gly-Pro-Arg-Gln-Ser-Asn-Glu-Thr-Pro-Gly-Ser-Pro-Ser-Gln-Glu-Glu-Arg (SEQ ID N 20), Gly-Pro-Arg-Gln-Ala-Lys-Thr-Leu-Pro-Gly-Ala-Pro-Ser-Gln-Thr-Thr-Arg (SEQ ID N 21) and Gly-Pro-Thr-Glu-Ala-Asp-Glu-Leu-Pro-Gly-Ala-Pro-Ser-Glu-Glu-Glu-Thr (SEQ ID N 22). 23. A superantigen conjugate of Claim 11, wherein x=0, y=1-3, and z=1-3. 24. The DNA molecule encoding a superantigen and immune modulator that is not a superantigen that is in the form of a biscistronic construct in which a first cistron contains a sequence encoding a superantigen, which is modified to have a decreased ability to bind to MRC class II antigen and decreased seroreactivity and immunogenicity in human blood serum compared to the corresponding wild type superantigen; and the other cistron contains a sequence encoding said immune modulator which is selected from the group consisting of cytokines, chemokines and extracellular parts of lymphocyte surface bound receptors and ligands. 25. The DNA molecule of Claim 24, wherein the immune modulator is IL-2. 26. The DNA molecule of Claim 24, characterized in that said encoded immune modulator is an extracellular part of a B7 molecule selected from the group, comprising CD80 and CD86. 27. The DNA molecule encoding modified superantigen thus it to have a decreased ability to bind to MRC class II antigen and decreased seroreactivity and immunogenicity in human blood serum compared to the corresponding wild type superantigen; immune modulator which is selected from the group consisting of cytokines, chemokines and extracellular parts of lymphocyte surface bound receptors and ligands, targeting moiety selected from the group, consisting of antibody, an antigen binding fragment thereof, Fab-fragment antibody or Fab2-fragment antibody or a single chain antibody. 28. The DNA molecule of Claim 27, wherein the immune modulator is IL-2. 29. The DNA molecule of Claim 27, wherein the immune modulator is an extracellular part of a B7 molecule is selected from the group, comprising CD80 and CD86. 30. A pharmaceutical composition for inactivating target cells comprising a superantigen, an immune modulator, and targeting moiety wherein at least one of the elements - superantigen or the immune modulator is conjugated with targeting moiety and targeting moiety is selected from the group, consisting of antibody, an antigen binding fragment thereof, Fab-fragment antibody or Fab2-fragment antibody or a single chain antibody, and said superantigen is modified to have a decreased ability to bind to MRC class II antigen and decreased seroreactivity and immunogenicity in human blood serum compared to the corresponding wild type superantigen; and said immune modulator is selected from the group consisting of cytokines, chemokines and extracellular parts of lymphocyte surface bound receptors and ligands, and said composition is taken in effective amount. 31 A pharmaceutical composition of Claim 30, characterized in that the superantigen is a chimera, comprising succession, derivative from two or more wild-type superantigens. 32. A pharmaceutical composition of Claim 30, characterized in that the immune modulator is IL-2. 33. A pharmaceutical composition of Claim 30, characterized in that the immune modulator is an extracellular part of a B7 molecule. 34. A pharmaceutical composition of Claim 33, characterized in that an extracellular part of a B7 molecule is selected from the group, comprising CD80 and CD86. 35. A pharmaceutical composition of Claim 30, characterized in that that the immune modulator has been modified to show a decreased affinity for its lymphocyte receptors, compared to the corresponding native form.