US 12,269,867 B2
Re-use of enzymes in in vitro glycoengineering of antibodies
Roberto Falkenstein, Penzberg (DE); Sebastian Malik, Penzberg (DE); Ingrid Grunert, Penzberg (DE); Marco Thomann, Penzberg (DE); Matthias Freiherr Von Roman, Penzberg (DE); and Heiko Walch, Penzberg (DE)
Assigned to Hoffmann-La Roche Inc., Little Falls, NJ (US)
Filed by Hoffmann-La Roche Inc., Little Falls, NJ (US)
Filed on Jun. 20, 2019, as Appl. No. 16/447,106.
Application 16/447,106 is a continuation of application No. PCT/EP2017/083430, filed on Dec. 19, 2017.
Claims priority of application No. 16205586 (EP), filed on Dec. 21, 2016; and application No. 17157005 (EP), filed on Feb. 20, 2017.
Prior Publication US 2020/0165321 A1, May 28, 2020
This patent is subject to a terminal disclaimer.
Int. Cl. C07K 16/00 (2006.01); C07K 1/18 (2006.01)
CPC C07K 16/00 (2013.01) [C07K 1/18 (2013.01); C07K 2317/21 (2013.01); C07K 2317/24 (2013.01); C07K 2317/31 (2013.01); C07K 2317/35 (2013.01); C07K 2317/41 (2013.01); C07K 2317/55 (2013.01)] 19 Claims
 
1. A method for the enzymatic production of an antibody with a modified glycosylation at an N-glycosylation site, comprising the following steps:
a) incubating a first batch of an antibody that has a glycosylation at an N-glycosylation site with one or more enzymes for a time sufficient and under conditions suitable to modify the glycosylation at the N-glycosylation site to a defined form,
b) separating in one or more chromatography steps the first batch of the antibody with a modified glycosylation at the N-glycosylation site from the one or more enzymes and thereby i) producing the antibody with a modified glycosylation at the N-glycosylation site and ii) obtaining one or more recycled enzymes, and
c) incubating a second batch of the antibody in the same production process that has a glycosylation at an N-glycosylation site with the one or more recycled enzymes of step b) for a time sufficient and under conditions suitable to modify the glycosylation at the N-glycosylation site to a defined form.