Enhancement of therapeutic protein in vivo activities through glycoengineering.
Delivery of protein therapeutics often requires frequent injections because of low activity or rapid clearance, thereby placing a burden on patients and caregivers. Using glycoengineering, we have increased and prolonged the activity of proteins, thus allowing reduced frequency of administration. Glycosylation analogs with new N-linked glycosylation consensus sequences introduced into the protein were screened for the presence of additional N-linked carbohydrates and retention of in vitro activity. Suitable consensus sequences were combined in one molecule, resulting in glycosylation analogs of rHuEPO, leptin, and Mpl ligand. All three molecules had substantially increased in vivo activity and prolonged duration of action. Because these proteins were of three different classes (rHuEPO is an N-linked glycoprotein, Mpl ligand an O-linked glycoprotein, and leptin contains no carbohydrate), glycoengineering may be generally applicable as a strategy for increasing the in vivo activity and duration of action of proteins. This strategy has been validated clinically for glycoengineered rHuEPO (darbopoetin alfa).
Elliott
Steve
S
Amgen, One Amgen Center, Thousand Oaks, CA 91320, USA. selliott@amgen.com
Lorenzini
Tony
T
Asher
Sheilah
S
Aoki
Ken
K
Brankow
David
D
Buck
Lynette
L
Busse
Leigh
L
Chang
David
D
Fuller
Janis
J
Grant
James
J
Hernday
Natasha
N
Hokum
Martha
M
Hu
Sylvia
S
Knudten
Andrew
A
Levin
Nancy
N
Komorowski
Renee
R
Martin
Frank
F
Navarro
Rachell
R
Osslund
Timothy
T
Rogers
Gary
G
Rogers
Norma
N
Trail
Geri
G
Egrie
Joan
J
D000740:Anemia / Q000188:drug therapy; D000818:Animals; D016466:CHO Cells / Q000378:metabolism; D019556:COS Cells / Q000378:metabolism; D006224:Cricetinae; D016503:Drug Delivery Systems / Q000379:methods*; D004921:Erythropoietin / Q000235:genetics / Q000378:metabolism / Q000627:therapeutic use; D005079:Excipients / Q000737:chemistry; D005260:Female; D024861:Genetic Enhancement / Q000379:methods; D006023:Glycoproteins / Q000096:biosynthesis* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D020738:Leptin / Q000096:biosynthesis / Q000172:deficiency / Q000235:genetics / Q000627:therapeutic use; D051379:Mice; D008807:Mice, Inbred BALB C; D015202:Protein Engineering / Q000379:methods*; D011506:Proteins / Q000008:administration & dosage / Q000627:therapeutic use*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000627:therapeutic use; D013926:Thrombopoietin / Q000096:biosynthesis / Q000627:therapeutic use
D005079:Excipients; D006023:Glycoproteins; D020738:Leptin; D011506:Proteins; D011994:Recombinant Proteins; D004921:Erythropoietin; D013926:Thrombopoietin