BISPECIFIC ANTIBODIES AGAINST CD3EPSILON AND ROR1
A bispecific antibody specifically binding to the two targets human CD3ε (further named also as “CD3”) and the extracellular domain of human ROR1 (further named also as “ROR1”), characterized in that the bispecific antibody does not internalize in a cell based assay at 37° C. during 2 hrs, using ROR1-positive B-CLL cells and used at an antibody concentration of 1 nM, whereby not internalize means, that the mean fluorescence intensity (MFI), as detected by flow cytometry, of a bispecific antibody upon binding to ROR1-positive primary B-CLL cells measured at time 0 is not reduced for more than 50%, preferably not more than 30% when re-measured after a 2 hr-incubation at 37° C. and which is useful for the treatment of B-cell malignancies like Chronic Lymphocytic Leukemia or B-cell disorders expressing ROR1 and ROR1-positive solid tumors.
1 . A trivalent bispecific antibody which specifically binds human CD3ε (CD3) and the extracellular domain of human ROR1 (ROR1), wherein the antibody comprises ROR1 Fab-Fc-CD3 Fab-ROR1 Fab ( FIG. 1 (1)).
2 . A trivalent bispecific antibody according to claim 1 , characterized in that the bispecific antibody does not internalize in a cell based assay at 37° C. during 2 hrs, using ROR1-positive primary B-CLL cells and used at an antibody concentration of 1 nM, whereby not internalize means, that the mean fluorescence intensity (MFI), as detected by flow cytometry, of said bispecific antibody upon binding to ROR1-positive primary B-CLL cells measured at time 0 is not reduced more than 50%, preferably not more than 30% when re-measured after a 2 hr-incubation at 37° C.
3 - 6 . (canceled)
7 . A trivalent bispecific antibody according to claim 1 , characterized in that the construct
consists of building blocks SEQ ID NO:30 (2×), 31, 32, and 33.
8 . A trivalent bispecific antibody according to claim 7 , characterized in that the anti-CD3ε antibody sequences VH and VL within SEQ ID NO: 31 and 33 are replaced by the respective VH and VL sequences of SEQ ID NO: 10 and 11.
9 - 10 . (canceled)
11 . A trivalent bispecific antibody according to claim 1 , characterized in that the variable domains VL and VH or the constant domains CL and CH1 of the anti-CD3 antibody are replaced by each other.
12 . A trivalent bispecific antibody according to claims 1 , characterized in that the antibody portion specifically binding to human CD3ε is characterized in comprising
a) a variable heavy chain domain VH comprising the CDRs of SEQ ID NO: 12, 13 and 14 as respectively heavy chain CDR1, CDR2 and CDR3 and a variable domain VL comprising the CDRs of SEQ ID NO: 15, 16 and 17 as respectively light chain CDR1, CDR2 and CDR3, or
b) a variable heavy chain domain VH comprising the CDRs of SEQ ID NO: 23, 24 and 25 as respectively heavy chain CDR1, CDR2 and CDR3 and a variable domain VL comprising the CDRs of SEQ ID NO: 26, 27 and 28 as respectively light chain CDR1, CDR2 and CDR3.
13 . A trivalent bispecific antibody according to claim 1 , characterized in that the antibody portion specifically binding to human ROR1 is characterized in comprising a variable heavy chain domain VH comprising the CDRs of SEQ ID NO: 7, 8 and 9 as respectively heavy chain CDR1, CDR2 and CDR3 and a variable domain VL comprising the CDRs of SEQ ID NO: 3, 4 and 5 as respectively light chain CDR1, CDR2 and CDR3.
14 - 15 . (canceled)
16 . A pharmaceutical composition comprising an antibody according to claim 1 and a pharmaceutically acceptable excipient.
17 - 19 . (canceled)