Modular antigen transporter molecules (MAT molecules) for modulating immune reactions, associated constructs, methods and uses

• US 20050281816A1
• Filed: 04/11/2005
• Published: 12/22/2005
• Est. Priority Date: 10/11/2002
• Status: Active Grant

• Current Assignee: Boehringer Ingelheim Vetmedica GmbH (C.H. Boehringer Sohn AG & Co. KG)
• Original Assignee: Imvision GmbH (ImVisioN Therapeutics AG)
• Inventors: Crameri, Reto, Fluckiger, Sabine, Daigle, Isabelle, Lamping, Norbert
• Granted Patent: US 7,563,866 B2
• Time in Patent Office: Days
• US Class Current: 424/144.1
• CPC Class Codes: A61K 39/00 Medicinal preparations containing antigens or antibodies materials for immunoassay G01N33/53
• CPC Class Codes: A61P 29/00 Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
• CPC Class Codes: A61P 31/00 Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
• CPC Class Codes: A61P 35/00 Antineoplastic agents
• CPC Class Codes: A61P 37/00 Drugs for immunological or allergic disorders
• CPC Class Codes: A61P 37/04 Immunostimulants
• CPC Class Codes: A61P 37/06 Immunosuppressants, e.g. drugs for graft rejection
• CPC Class Codes: A61P 37/08 Antiallergic agents antiasthmatic agents A61P11/06; ophthalmic antiallergics A61P27/14
• CPC Class Codes: A61P 39/00 General protective or antinoxious agents
• CPC Class Codes: C07K 14/38 from Aspergillus
• CPC Class Codes: C07K 2319/10 containing a tag for extracellular membrane crossing, e.g. TAT or VP22
• CPC Class Codes: C07K 2319/40 containing a tag for immunodetection, or an epitope for immunisation
• CPC Class Codes: C12N 15/62 DNA sequences coding for fusion proteins
• CPC Class Codes: C12N 2310/111 spanning the whole gene, or a large part of it
• CPC Class Codes: C12N 2810/40 Vectors comprising a peptide as targeting moiety, e.g. a synthetic peptide, from undefined source

Abstract

It is therefore the object of the invention to provide means for supplying any of a wide variety of antigens in a sufficiently targeted manner to their processing site that even better therapeutic and diagnostic use thereof is possible.

Claims

1. A modular antigen transport molecule (MAT molecule) comprising at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecule from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecule intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecule in an individual.

2. The MAT molecule as claimed in claim 1, in which the at least one translocation module comprises at least one of the following sequences, or an amino acid sequence that is at least 90% identical to one of the following sequences, or at least one functional partial sequence of these sequences:

HIV tat, antennapedia, VP22, polyarginine or polylysine sequences having a length of from 4 to 20 amino acids, or sequences which are inverted relative to the aforementioned, where the module may be increased or reduced in its translocation efficiency through modification of its sequence, and where the translocation module is present N-terminally, C-terminally or internally in the MAT molecule.

3. The MAT molecule as claimed in claim 1, in which the at least one targeting module comprises one of the following sequences or structures, or an amino acid sequence that is at least 90% identical to one of the following sequences, or at least one functional partial sequence of these sequences:

major histocompatibility complex (MHC) invariant chain, MHC II beta chain, mannose 6-phosphate receptor, membrane-bound immunoglobulin (mIg), HLA-DM beta chain, LAMP-1, LAMP-2, LAMP-3, Limp 1, CD63, CD82, CD1b or sequences which are inverted relative to the aforementioned, or a mannose 6-phosphate structure, where the module is increased or reduced in its targeting efficiency preferably by modification of its sequence, and where the targeting module is present N-terminally, C-terminally or internally in the MAT molecule.

4. The MAT molecule as claimed in claim 1, in which the at least one antigen module is a protein, a peptide, a nucleic acid, a sugar structure, a lipid, or another antigen, and where the antigen module is present N-terminally, C-terminally or internally in the MAT molecule, or is introduced internally into a targeting module, a translocation module or a tag module, and where the antigen can be produced by use of combinatorial methods.

5. The MAT molecule as claimed in claim 4, in which the at least one antigen module is a protein or peptide derived from a complete Fel d1 allergen or fragments of the Fel d 1 allergen, or a protein at least 90% identical to Fel d 1 allergen.

6. The MAT molecule as claimed in claim 4, in which the at least one antigen module is a protein or peptide derived from a Bet v 1 or Api g 1 allergen or fragments of the Bet v 1 or Api g 1 allergen or a protein at least 90% identical to Bet v 1 or Api g 1 allergen.

7. The MAT molecule as claimed in claim 4, in which the at least one antigen module is a protein or peptide derived from a complete Der p 1 allergen or fragments of the Der p 1 allergen or a protein at least 90% identical to Der p 1 allergen.

8. The MAT molecule as claimed in claim 4, in which the at least one antigen module is a protein or peptide derived from a complete PLA2 allergen or fragments of the PLA2 allergen, or a protein at least 90% identical to PLA2 allergen.

11. The MAT molecule as claimed in claim 1, in which the individual modules are coupled to one another by covalent and/or noncovalent linkages, where noncovalent linkages may be brought about by one or more of the following interactions:

a) protein/protein interactions

b) nucleic acid/protein interactions

c) nucleic acid/nucleic acid interactions

d) sugar/lectin interactions

e) receptor/ligand interactions

f) antigen/antibody interactions

g) hapten/antibody interactions

h) Spiegelmer/interactions

i) streptavidin/biotin interactions

j) avidin/biotin interactions

k) protein A/antibody interactions

l) protein G/antibody interactions

m) protein L/antibody interactions

n) histidine peptide/nickel chelate interactions

o) glutathione/glutathione S-transferase interactions

p) maltose/maltose-binding protein interactions and

q) chitin/chitin-binding protein interactions

and where covalently connected modules may be connected together via a spacer module.

12. The MAT molecule as claimed in claim 1, which comprises at least one of the following building blocks:

a) D isomers of amino acids

b) modified D isomers of amino acids

c) L isomers of amino acids

d) modified L isomers of amino acids

e) building blocks having guanidino groups

f) building blocks having amidino groups

g) peptidomimetics

h) N-methyl amino acids

i) unusual amino acids and

j) Spiegelmers

14. The MAT molecule as claimed in claim 1, in which one or more of the following are present:

a) a translocation module in more than one copy

b) a targeting module in more than one copy

c) an antigen module in more than one copy

d) a tag module in more than one copy and

e) a spacer module in more than one copy.

15. The MAT molecule as claimed in claim 1, which is composed wholly or partly of one or more of cyclic, branched and dendrimeric arrangements of modules.

16. The Mat molecule as claimed in claim 1, wherein the translocation module comprises the HIV-tat sequence or a fragment thereof, the targeting module comprises the human invariant chain or a fragment thereof, and the antigen module comprises a fusion-peptide of Fel d 1 chain 2 fused to the N-terminus of Fel d 1 chain 1, or a fragment of the fusion-peptide or a fragment of one or both chains of Fel d 1.

17. The MAT molecule as claimed in claim 16 having the sequence as depicted in a SEQ ID selected from the group consisting of SEQ ID NO:

21, SEQ ID NO;

22, SEQ ID NO;

23, SEQ ID NO;

24, SEQ ID NO;

25, SEQ ID NO;

26, SEQ ID NO;

27, SEQ ID NO;

28, SEQ ID NO;

29, SEQ ID NO;

30, SEQ ID NO;

31, SEQ ID NO;

32, SEQ ID NO;

33, SEQ ID NO;

34, SEQ ID NO;

35 and SEQ ID NO;

36.

9-10. -10. (canceled)

13. (canceled)

18. (canceled)

19. A nucleic acid which codes for MAT molecules wherein said MAT molecules comprise at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecule from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecule intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecule in an individual.

20-29. -29. (canceled)

30. A medicament, vaccine or diagnostic aid comprising MAT molecules wherein said MAT molecules comprise at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecules from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecules intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecules in an individual.

31. The medicament, vaccine or diagnostic aid according to claim 30, further comprising at least one of the following constituents:

a) a physiological sodium chloride solution

b) a pharmaceutically acceptable adjuvant

c) a pharmaceutically acceptable buffer substance

d) a pharmaceutically acceptable carrier protein

e) a pharmaceutically acceptable preservative and

f) a pharmaceutically acceptable colorant.

32. The medicament, vaccine or diagnostic aid as claimed in claim 30, characterized in that the product is designed for subcutaneous injection, injection into a lymph node or for administration via mucous membranes.

33. The medicament, vaccine or diagnostic aid as claimed in claim 30, characterized in that it is intended for one or more uses selected from the group consisting of

a) prophylaxis

b) reducing or enhancing an immune reaction

c) treatment of a condition selected from the group consisting of an autoimmune disease, an allergy, a rheumatic disorder, a rejection reaction against a transplanted organ, an infectious disease, induced by viral, bacterial or eukaryotic pathogens, and a malignant disease.

34-38. -38. (canceled)

39. A method for treating or diagnosing a malignant disease, an allergy, or an infectious disease in an individual, comprising the step of providing said individual with a sufficient amount of a MAT-molecule or a composition comprising a MAT-molecule, wherein said MAT molecule comprises at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecule from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecule intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecule in an individual.

40-41. -41. (canceled)

42. A method for treatment of a disease comprising the step of injecting MAT-molecules or a composition comprising MAT-molecules into a lymph node.

43. A method for producing MAT molecules comprising the steps of

preparing and characterizing individual modules of the MAT molecules;

selecting at least one targeting module;

selecting at least one translocation module;

selecting at least one antigen module; and

combining said at least one targeting module, said at least one translocation module and said at least one antigen module to give a noncovalently or covalently connected MAT molecule, wherein said step of combining is carried out on the basis of one or more of the following;

molecular properties of the individual modules, affinities of the individual modules for one another, reactive chemical groups of the individual modules, and chosen reaction conditions.

44. A cell, primary cell or cell line in which antigen presentation has been modulated, wherein said cell comprises i) at least one MAT molecule comprising at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecule from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecule intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecule in an individual;

or

ii) at least one nucleic acid which codes for said at least one MAT molecule.

45. The cell, primary cell or cell line of claim 44, wherein said at least one nucleic acid is associated with a vector.

46. A medicament, vaccine or diagnostic aid comprising a nucleic acid which codes for MAT molecules wherein said MAT molecules comprise at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecule from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecule intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecule in an individual.

47. A medicament, vaccine or diagnostic aid comprising cells which comprise at least one nucleic acid sequence which codes for a MAT molecule, wherein said MAT molecule comprises at least one translocation module, at least one targeting module and at least one antigen molecule, where

a) the translocation module is a molecular unit which brings about transport of the MAT molecule from the extracellular space into the interior of the cell,

b) the targeting module is a molecular unit which brings about transport of the MAT molecule intracellularly to the organelles which are involved in the processing of antigens or the loading of major histocompatibility complex (MHC) molecules with antigens, and

c) the antigen module is a molecular unit which at least consists of one antigen and which determines the specificity of an immune response modulated by the MAT molecule in an individual.