IMMUNO-BASED RETARGETED ENDOPEPTIDASE ACTIVITY ASSAYS
First Claim
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1. A method of detecting retargeted endopeptidase activity, the method comprising the steps of:
- a) treating a cell from an established cell line with a sample comprising a retargeted endopeptidase, wherein the cell from an established cell line is susceptible to retargeted endopeptidase activity by a retargeted endopeptidase;
b) isolating from the treated cell line a SNAP-25 component comprising a SNAP-25 cleavage product having a carboxyl-terminus at the P1 residue of the BoNT/A cleavage site scissile bond;
c) contacting the SNAP-25 component with an α
-SNAP-25 antibody, wherein the α
-SNAP-25 antibody binds an epitope comprising a carboxyl-terminus at the P1 residue of the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product; and
d) detecting the presence of an antibody-antigen complex comprising the α
-SNAP-25 antibody and the SNAP-25 cleavage product;
wherein detection by the antibody-antigen complex is indicative of retargeted endopeptidase activity, wherein the α
-SNAP-25 antibody binds an epitope comprising a carboxyl-terminus at the P1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product and wherein the isolated α
-SNAP-25 antibody has a VH CDR1 of SEQ ID NO;
93, a VH CDR2 of SEQ ID NO;
96, a VH CDR3 of SEQ ID NO;
100, a VL CDR1 of SEQ ID NO;
105, a VL CDR2 of SEQ ID NO;
110, and a VL CDR3 of SEQ ID NO;
115.
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Abstract
The present specification discloses a retargeted endopeptidase pharmaceutical wherein the activity has been determined by the methods disclosed.
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Citations
10 Claims
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1. A method of detecting retargeted endopeptidase activity, the method comprising the steps of:
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a) treating a cell from an established cell line with a sample comprising a retargeted endopeptidase, wherein the cell from an established cell line is susceptible to retargeted endopeptidase activity by a retargeted endopeptidase; b) isolating from the treated cell line a SNAP-25 component comprising a SNAP-25 cleavage product having a carboxyl-terminus at the P1 residue of the BoNT/A cleavage site scissile bond; c) contacting the SNAP-25 component with an α
-SNAP-25 antibody, wherein the α
-SNAP-25 antibody binds an epitope comprising a carboxyl-terminus at the P1 residue of the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product; andd) detecting the presence of an antibody-antigen complex comprising the α
-SNAP-25 antibody and the SNAP-25 cleavage product;
wherein detection by the antibody-antigen complex is indicative of retargeted endopeptidase activity, wherein the α
-SNAP-25 antibody binds an epitope comprising a carboxyl-terminus at the P1 residue from the BoNT/A cleavage site scissile bond from a SNAP-25 cleavage product and wherein the isolated α
-SNAP-25 antibody has a VH CDR1 of SEQ ID NO;
93, a VH CDR2 of SEQ ID NO;
96, a VH CDR3 of SEQ ID NO;
100, a VL CDR1 of SEQ ID NO;
105, a VL CDR2 of SEQ ID NO;
110, and a VL CDR3 of SEQ ID NO;
115. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9)
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10. An isolated monoclonal α
- -SNAP-25 antibody which has a VH CDR1 of SEQ ID NO;
93, a VH CDR2 of SEQ ID NO;
96, a VH CDR3 of SEQ ID NO;
100, a VL CDR1 of SEQ ID NO;
105, a VL CDR2 of SEQ ID NO;
110, and a VL CDR3 of SEQ ID NO;
115.
- -SNAP-25 antibody which has a VH CDR1 of SEQ ID NO;
Specification