Bispecific antigen binding molecules comprising lipocalin muteins
An antigen-binding molecule, lipocalin technology, applied in immunoglobulin, anti-enzyme immunoglobulin, hybrid immunoglobulin, etc., can solve the problem of patients not benefiting
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[0396] The following are examples of methods and compositions of the invention. It is understood that various other embodiments may be practiced, given the general description provided above.
[0397] recombinant DNA technology
[0398] DNA was manipulated using standard methods, as described in Sambrook et al., Molecular cloning: A laboratory manual; Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, 1989. Molecular biology reagents were used according to the manufacturer's instructions. General information on the nucleotide sequences of human immunoglobulin light and heavy chains is given in the following reference: Kabat, E.A. et al., (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, NIH Publication No 91- 3242.
[0399] DNA sequencing
[0400] DNA sequence was determined by double-strand sequencing.
[0401] gene synthesis
[0402] Desired gene segments were generated by PCR using appropriate templates or by automated gene synthes...
example 1
[0412] Preparation, purification and characterization of bispecific antibodies with bivalent binding to 4-1BB and monovalent / bivalent binding to FAP
[0413] 1.1 Generation of bispecific antibodies with bivalent binding to 4-1BB and monovalent or bivalent binding to FAP
[0414] The preparation method of bispecific agonistic 4-1BB antibody with bivalent binding to 4-1BB and monovalent or bivalent binding to FAP is as follows Figure 1A with 1B shown. FAP conjugate (clone 4B9, produced and prepared as described in WO 2012 / 020006 A2, which is incorporated herein by reference) and 4-1BB conjugate (anticalin, produced and prepared as described in WO 2016 / 177802) with in preparation Figure 1A with 1B Molecules described in wherein TA1 is FAP. According to the method described in International Patent Application Publication No. WO2012 / 130831A1, Pro329Gly, Leu234Ala, and Leu235Ala mutations were introduced into the Fc constant region of the heavy chain to abolish binding to Fcγ...
example 2
[0434] Preparation, purification and characterization of bispecific antibodies with bivalent binding to 4-1BB and monovalent / bivalent binding to HER2
[0435] 2.1 Generation of bispecific antibodies with bivalent binding to 4-1BB and monovalent or bivalent binding to HER2
[0436] The preparation method of bispecific agonistic 4-1BB antibody with bivalent binding to 4-1BB and monovalent or bivalent binding to HER2 is as follows: Figure 1A and 1B shown. HER2 conjugates (corresponding to trastuzumab) and 4-1BB conjugates (lipocalin, produced and prepared as described in WO 2016 / 177802) were used in the preparation Figure 1A and 1B Molecules described in wherein TA1 is HER2. According to the method described in International Patent Application Publication No. WO2012 / 130831A1, Pro329Gly, Leu234Ala, and Leu235Ala mutations were introduced into the Fc constant region of the heavy chain to abolish binding to Fcγ receptors.
[0437] The variable regions of the heavy and light c...
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