Insights into the Impact of Heterogeneous Glycosylation on the Pharmacokinetic Behavior of Follistatin-Fc-Based Biotherapeutics.
Follistatin 315 heparan sulfate-binding deficient mutant human IgG4 Fc fusion (FST-ΔHBS-Fc) is a follistatin (FST) based Fc fusion protein currently being developed as a novel therapy for several potential indications, including muscle wasting. Previous assessments of the pharmacokinetics and therapeutic activity of FST-ΔHBS-Fc have shown a close association of the exposure-response relationship. The current work builds upon these initial studies by investigating the glycosylation characteristics of FST-ΔHBS-Fc after recombinant expression and its impact on the pharmacokinetics in mice and Cynomolgus monkeys. The data presented indicate that FST-ΔHBS-Fc is heterogeneously glycosylated at the three putative sites in FST when recombinantly expressed in stably transfected Chinese hamster ovary cells. Such carbohydrate heterogeneity, especially with regards to sialic acid incorporation, directly results in sugar-dependent clearance in both mice and Cynomolgus monkeys. Examination of the pharmacokinetics of FST-ΔHBS-Fc molecules containing variable sialic acid content in asialoglycoprotein receptor 1 (ASPGR-1) knockout mice supports the receptor's role as part of the clearance mechanism of the molecules. Based on the evaluation of several variably sialylated lots of material in pharmacokinetic assessments, we define specifications for average sialic acid incorporation into FST-ΔHBS-Fc that result in limited sugar-mediated clearance. Taken together, these studies highlight the importance of establishing an early understanding of the glycosylation/pharmacokinetic relationships of FST-ΔHBS-Fc, which will provide a basis for future application toward optimal systemic drug delivery and dosing strategies.
Departments of Drug Disposition Development/Commercialization (A.D.-M.), Biotechnology Discovery Research (J.P., J.E.C), Bioproduct Research and Development (L.H.), and Musculoskeletal Research (B.Y.), Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana; Biotechnology Discovery Research, Applied Molecular Evolution, San Diego, California (A.K.).
Amita
A
Datta-Mannan
Departments of Drug Disposition Development/Commercialization (A.D.-M.), Biotechnology Discovery Research (J.P., J.E.C), Bioproduct Research and Development (L.H.), and Musculoskeletal Research (B.Y.), Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana; Biotechnology Discovery Research, Applied Molecular Evolution, San Diego, California (A.K.).
Lihua
L
Huang
Departments of Drug Disposition Development/Commercialization (A.D.-M.), Biotechnology Discovery Research (J.P., J.E.C), Bioproduct Research and Development (L.H.), and Musculoskeletal Research (B.Y.), Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana; Biotechnology Discovery Research, Applied Molecular Evolution, San Diego, California (A.K.).
Jennifer
J
Pereira
Departments of Drug Disposition Development/Commercialization (A.D.-M.), Biotechnology Discovery Research (J.P., J.E.C), Bioproduct Research and Development (L.H.), and Musculoskeletal Research (B.Y.), Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana; Biotechnology Discovery Research, Applied Molecular Evolution, San Diego, California (A.K.).
Benjamin
B
Yaden
Departments of Drug Disposition Development/Commercialization (A.D.-M.), Biotechnology Discovery Research (J.P., J.E.C), Bioproduct Research and Development (L.H.), and Musculoskeletal Research (B.Y.), Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana; Biotechnology Discovery Research, Applied Molecular Evolution, San Diego, California (A.K.).
Andrew
A
Korytko
Departments of Drug Disposition Development/Commercialization (A.D.-M.), Biotechnology Discovery Research (J.P., J.E.C), Bioproduct Research and Development (L.H.), and Musculoskeletal Research (B.Y.), Lilly Research Laboratories, Lilly Corporate Center, Indianapolis, Indiana; Biotechnology Discovery Research, Applied Molecular Evolution, San Diego, California (A.K.) croy_johnny_eugene@lilly.com.
Johnny E
JE
Croy
D000818:Animals; D001691:Biological Therapy / Q000639:trends*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D038681:Follistatin / Q000737:chemistry* / Q000493:pharmacokinetics*; D006031:Glycosylation; D057809:HEK293 Cells; D006801:Humans; D008252:Macaca fascicularis; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D016513:Mice, SCID
D038681:Follistatin