Recombinant human mannan-binding lectin
OwnerSteffen Thiel, Jens Christian Jensenius +1inventors
InventorsSteffen Thiel, Jens Christian Jensenius, Thomas Vorup Jensen
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Abstract
The present invention relates to methods of producing and/or isolating MBL compositions. In particular the invention relates to methods of producing and/or isolating novel recombinant human MBL preparations with high similarity to natural human MBL. The MBL preparations may be used in compositions, medicaments and in methods for treatment of conditions such as those related to immunosuppressive conditions and/or to conditions of MBL deficiencies including latent conditions. Conditions of deficiencies of Mannan-Binding Lectin (MBL) are associated with increased susceptibility to infections.
Claims30
A process of producing a purified recombinant human mannan binding lectin (MBL) composition which is free of monomers of the 96 kDa MBL subunit which comprises
- providing a starting recombinant human MBL composition comprising both (a) monomers of the 96 kDa MBL subunit and (b) at least one oligomer selected from the group consisting of dimers, trimers, tetramers, pentamers and hexamers of said 96 kDa MBL subunit, wherein said 96 kDa subunit comprises the polypeptide of SEQ ID NO:1
- separating (b) from (a) by subjecting the composition to affinity chromatography using a carbohydrate-derivatized matrix having affinity for at least one oligomer of (b) and which substantially lacks affinity for said monomers of (a), and thereby obtaining a purified composition which comprises at least one oligomer of (b) and which is without any detectable amount of said monomer (a) of said 96 kDa MBL subunit.
A process of producing a purified recombinant human mannan binding lectin (MBL) compositions which is enriched for at least one higher oligomer selected from the group consisting of tetramers, pentamers and hexamers of the 96 kDa MBL subunits, which comprises
- providing a starting recombinant human MBL composition comprising both (a) monomers of the 96 kDa MBL subunit and (b) at least one higher oligomer selected from the group consisting of tetramers, pentamers and hexamers of said subunit,
- separating (b) from (a) by subjecting the composition to affinity chromatography using a carbohydrate-derivatized matrix which has a higher affinity for (b) than for (a), thereby obtaining a purified composition wherein the ratio of (1) the sum of the tetramers, pentamers and hexamers of the 96 kDa MBL subunit, to (2) the monomers of the 96 kDa MBL subunit, is at least 5:1.
A process of producing a purified recombinant human mannan binding lectin (MBL) composition, which is enriched for at least one higher oligomer selected from the group consisting of tetramers, pentamers and hexamers of the 96 kDa MBL subunits, which comprises
- providing a starting recombinant human MBL composition comprising both (a) monomers and/or dimers of the 96 kDa MBL subunit and (b) at least one higher oligomer selected from the group consisting of, tetramers, pentamers and hexamers of said subunit,
- separating (b) from (a) by subjecting the composition to affinity chromatography using a carbohydrate-derivatized matrix which has a higher affinity for (b) than for (a), thereby obtaining a purified composition wherein the ratio of (1) the sum of the tetramers, pentamers and hexamers of the 96 kDa MBL subunit, to (2) the sum of the monomers and dimers of the 96 kDa MBL subunit, is at least 2:1.
A process of producing a purified recombinant human mannan binding lectin (MBL) composition which is enriched for hither oligomers, selected from the group consisting of tetramers, pentamers, and hexamers of the 96 kDa MBL subunit, which comprises
- providing a starting recombinant human MBL composition which comprises (a) monomers and/or dimers of the 96 kDa MBL subunit, and (b) said higher oligomers, of said subunit,
- subjecting the starting composition to affinity chromatography using a carbohydrate-derivatized matrix which has a higher affinity for at least one of said oligomers of (b) than for said monomers and/or dimers thereof, thereby obtaining a purified composition enriched, relative to said starting composition, for said oligomers of (b), and in which more than 50% of the MBL is in the form of said oligomers of (b).
The process of 15 in which more than 60% of the MBL in the purified composition is in the form of said oligomers of (b).
The process of 15 in which more than 70% of the MBL in the purified composition is in the form of said oligomers of (b).
The process of 15 in which more than 80% of the MBL in the purified composition is in the form of said oligomers of (b).
The process of 15 in which more than 90% of the MBL in the purified composition is in the form of said oligomers of (b).
The process of 15 in which more than 95% of the MBL in the purified composition is in the form of said oligomers of (b).
A process of producing a purified recombinant human mannan binding lectin (MBL) composition which is free of monomers of the 96 kDa MBL subunit which comprises
- providing a starting recombinant human MBL composition comprising both (a) monomers of the 96 kDa MBL subunit and (b) at least one oligomer selected from the group consisting of dimers, trimers, tetramers, pentamers and hexamers of said 96 kDa MBL subunit,
- separating (b) from (a) by subjecting the composition to affinity chromatography using a carbohydrate-derivatized matrix having affinity for at least one oligomer of (b) and which substantially lacks affinity for said monomers of (a), and thereby obtaining a purified composition which comprises at least one oligomer of (b) and which is without any detectable amount of said monomer (a) of said 96 kDa MBL subunit
- wherein said starting recombinant MBL composition is obtained by
- providing a nonhuman transgenic mammal whose cells comprise an expression construct encoding the human MBL polypeptide of SEQ ID NO:1,
- producing said polypeptide in said mammal,
- and recovering said starting MBL composition from at least one body tissue or body fluid of said mammal.
Dependencies are inferred from the claim text.