Antibodies to MASP-2
View Patent ↗The invention relates to antibodies to MASP-2 and functional equivalents thereof. In particular, the invention relates to MASP-2 antibodies capable of inhibiting the function of MASP-2. The invention furthermore discloses MASP-2 epitopes, wherein antibodies recognizing said epitopes are in particularly useful for inhibiting MASP-2 activity. The invention also relates to methods of producing said antibodies, methods of inhibiting MASP-2 activity as well as to pharmaceutical compositions comprising the MASP-2 antibodies.
1. A method of producing an antibody specifically recognizing and binding an epitope within the C-terminal part of human MASP-2 comprising the CCP1, CCP2 and serine protease domains, wherein said antibody is capable of inhibiting deposition of C4 substrate added to MBL/MASP-2 complexes present in full human serum obtained from individuals with C4 deposition activity, said method comprising the steps of
(a) identifying one or more test antibodies capable of binding an epitope within a polypeptide fragment of human MASP-2 consisting of the CCP1, CCP2 and serine protease domains (aa 293 to 686 of SEQ ID NO:1, comprising:
(i) providing human MASP-2 or a polypeptide fragment of human MASP-2 consisting of the CCP1, CCP2 and serine protease domains (aa 293 to 686 of SEQ ID NO:1);
(ii) adding a test antibody and a reference antibody to the said MASP-2, wherein either the test antibody or the reference antibody is labeled with a detectable label or both antibodies are labeled with different detectable labels; wherein the reference antibody is selected from the group consisting of:
1) the monoclonal antibody produced by the hybridoma cell line deposited under the deposit number 03050904;
2) the monoclonal antibody produced by the hybridoma cell line deposited under the deposit number DSM ACC2657;
3) the monoclonal antibody produced by the hybridoma cell line deposited under the deposit number DSM ACC26660;
4) the monoclonal antibody produced by the hybridoma cell line deposited under the deposit number DSM ACC2658; and
5) the monoclonal antibody produced by the hybridoma cell line deposited under the deposit number DSM ACC2659;
iii) detecting the presence of the detectable label at MASP-2, thereby detecting whether the test antibody is capable of displacing the reference antibody;
iv) selecting test antibodies capable of displacing the reference antibody;
(b) testing whether antibodies identified in accordance with step (a) are capable of inhibiting deposition of C4 substrate added to MBL/MASP-2 complexes present in full human serum obtained from individuals with C4 deposition activity and
(c) selecting antibodies capable of inhibiting C4 deposition in accordance with step (b).
2. The method according to claim 1 , wherein the one or more test antibodies identified in accordance with step (a) are obtained from a mammalian subject after administration of a polypeptide fragment of human MASP-2 consisting of the CCP1, CCP2 and serine protease domains (aa 293 to 686 of SEQ ID NO:1), the method furthermore comprises isolating antibody producing cells from said mammal, preparing hybridoma cells from said antibody producing cells, cultivating said hybridomas and isolating antibodies produced by said hybridomas.
3. The method of claim 1 , comprising selecting antibodies capable of inhibiting deposition of C4 substrate added to MBL/MASP-2 complexes present in full human serum to less than 30% of control C4 deposition.
4. The method of claim 1 , comprising selecting antibodies capable of inhibiting deposition of C4 substrate added to MBL/MASP-2 complexes present in full human serum to less than 50% of control C4 deposition.
5. The method of claim 1 , wherein the one or more test antibodies identified in accordance with step (a) are recombinant antibodies.
6. The method of claim 5 , wherein the recombinant antibodies are produced using phage display.