Methods for purifying heterodimeric multispecific antibodies from parental homodimeric antibody species

Abstract

Methods for purifying multispecific antibodies on interest (MAIs) that co-engage at least two different antigens or epitopes (also referred to targets, used interchangeably throughout), from compositions comprising the MAI and parental homodimeric antibody species are provided, as well as reagents which may be used to practice such methods.

1 - 59 . (canceled) 
     
     
         60 . An ion exchange eluant comprising either:
 (i) CAPS, CHES, TAPS, HEPPSO, MOPSO, MES, acetic acid, formic acid, and a salt; or   (ii) methylamine, 1,2-ethanediamine, 1-methylpiperazine, 1,4-dimethylpiperazine, 2-[Bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)propane-1,3-diol (bis-tris), and hydroxylamine and optionally at least one salt.   
     
     
         61 . (canceled) 
     
     
         62 . The ion exchange eluant according to  claim 60 , with the proviso that the eluant does not include TRIS, piperazine, or imidazole. 
     
     
         63 . (canceled) 
     
     
         64 . (canceled) 
     
     
         65 . The ion exchange eluant according  claim 60 , wherein the salt is selected from the group consisting of NaCl, KCl, or Na 2 SO 4 . 
     
     
         66 . The ion exchange eluant according  claim 60 , wherein the eluant is suitable for use in the purification of an MAI from a composition comprising the MAI and parental homodimeric antibody species. 
     
     
         67 . The ion exchange eluant according to  claim 60 , wherein the eluant comprises at least one salt at a concentration range selected from the group consisting of: 0 mM to about 100 mM; 0 mM to about 60 mM; 0 mM to about 50 mM; 0 mM to about 40 mM; 0 mM to about 30 mM; 0 mM to about 20 mM; 0 mM to about 10 mM; 0 mM to about 5 mM; about 10 mM to about 200 mM; about 10 mM to about 100 mM; about 10 mM to about 50 mM; about 10 mM to about 40 mM; about 10 mM to about 30 mM; about 10 mM to about 20 mM; about 20 mM to about 200 mM; about 20 mM to about 100 mM; about 20 mM to about 50 mM; about 20 mM to about 30 mM; about 30 mM to about 200 mM; about 30 mM to about 100 mM; and about 30 mM to about 50 mM; and about 5 mM to about 15 mM. 
     
     
         68 . The ion exchange eluant according to  claim 60 , wherein each sample of the eluant comprises at least one salt at a concentration of about 10 mM. 
     
     
         69 . The ion exchange eluant according to  claim 60 , wherein the salt is NaCl. 
     
     
         70 . The ion exchange eluant according to  claim 60 , wherein each sample of the eluant comprises NaCl at a concentration of about 10 mM. 
     
     
         71 . The ion exchange eluant according to  claim 60 , wherein the ion exchange eluant comprises one or more antibodies. 
     
     
         72 . The ion exchange eluant according to  claim 71 , wherein the one or more antibodies comprise either a first parental homodimeric antibody species or a second parental homodimeric antibody species. 
     
     
         73 . The ion exchange eluant according to  claim 72 , wherein the difference in isoelectric point (pI) between the first and second parental homodimeric antibody species is less than 0.5 pH units. 
     
     
         74 . The ion exchange eluant according to  claim 73 , wherein the different isoelectric points are actual isoelectric points or calculated isoelectric points and either:
 a. the difference in the actual isoelectric point of the first heavy chain polypeptide and the actual isoelectric point of the second heavy chain polypeptide is less than 0.50 pH unit, less than 0.45 pH unit; less than 0.40 pH unit; less than 0.35 pH unit; less than 0.30 pH unit; less than 0.25 pH unit; less than 0.20 pH unit; less than 0.15 pH unit; less than 0.14 pH unit; less than 0.13 pH unit; less than 0.12 pH unit; less than 0.11 pH unit; less than 0.10 pH unit; less than 0.09 pH unit; less than 0.08 pH unit; less than 0.07 pH unit; less than 0.06 pH unit less than 0.04 pH unit; less than 0.03 pH unit; less than 0.025 pH unit; less than 0.02 pH unit; or pH values that are between any of the preceding values;   b. the difference in the actual isoelectric point of the first parental homodimeric antibody species and the actual isoelectric point of the second parental homodimeric antibody species is less than 0.50 pH unit; less than 0.45 pH unit; less than 0.40 pH unit; less than 0.35 pH unit; less than 0.30 pH unit; less than 0.25 pH unit; less than 0.20 pH unit; less than 0.15 pH unit; less than 0.14 pH unit; less than 0.13 pH unit; less than 0.12 pH unit; less than 0.11 pH unit; less than 0.10 pH unit; less than 0.09 pH unit; less than 0.08 pH unit; less than 0.07 pH unit; less than 0.06 pH unit less than 0.04 pH unit; less than 0.03 pH unit; less than 0.025 pH unit; less than 0.02 pH unit; or pH values that are between any of the preceding values;   c. the difference in the calculated isoelectric point of the first heavy chain polypeptide and the calculated isoelectric point of the second heavy chain polypeptide is less than 0.50 pH unit; less than 0.45 pH unit; less than 0.40 pH unit; less than 0.35 pH unit; less than 0.30 pH unit; less than 0.25 pH unit; less than 0.20 pH unit; less than 0.15 pH unit; less than 0.14 pH unit; less than 0.13 pH unit; less than 0.12 pH unit; less than 0.11 pH unit; less than 0.10 pH unit; less than 0.09 pH unit; less than 0.08 pH unit; less than 0.07 pH unit; less than 0.06 pH unit less than 0.04 pH unit; less than 0.03 pH unit; less than 0.025 pH unit; less than 0.02 pH unit; or pH values that are between any of the preceding values; or   d. the difference in the calculated isoelectric point of the first parental homodimeric antibody species and the calculated isoelectric point of the second parental homodimeric antibody species is less than 0.50 pH unit; less than 0.45 pH unit; less than 0.40 pH unit; less than 0.35 pH unit; less than 0.30 pH unit; less than 0.25 pH unit; less than 0.20 pH unit; less than 0.15 pH unit; less than 0.14 pH unit; less than 0.13 pH unit; less than 0.12 pH unit; less than 0.11 pH unit; less than 0.10 pH unit; less than 0.09 pH unit; less than 0.08 pH unit; less than 0.07 pH unit; less than 0.06 pH unit less than 0.04 pH unit; less than 0.03 pH unit; less than 0.025 pH unit; less than 0.02 pH unit; or pH values that are between any of the preceding values.