The numerical labels for each species are as described in the text. are shown. Verinurad Within the adipose, the intact and N-only forms can bind to the receptor complex with indicated binding affinities.(TIF) pone.0119104.s001.tif (1.8M) GUID:?796C2F04-00F6-4C94-9266-F2935E33E2E1 Data Availability StatementAll relevant data are within the paper and its Supporting Information files. Abstract Pharmacological administration of fibroblast growth factor 21 (FGF21) improves metabolic profile in preclinical species and humans. FGF21 exerts its metabolic effects through formation of beta-klotho (KLB)/FGF receptor 1c FGFR1c complex and subsequent signaling. Data from various systems demonstrate the intact C- and N-terminus of FGF21 is required for binding with KLB, and interaction with FGFR1c, respectively. However the relative roles of the termini for pharmacological effects are unclear. Here we report PF-05231023, a long-acting FGF21 analogue which is unique in that the half-life and subcutaneous (SC) bioavailability of the intact C-terminus are significantly different from those of the intact N-terminus (2 vs. 22 hr Verinurad for half-life and 4~7 vs. ~50% SC bioavailability). Therefore, this molecule serves as a valuable tool to evaluate the relative roles of intact C-terminus vs. N-terminus in pharmacology studies in preclinical species. We determined the effects of PF-05231023 administration on body weight (BW) loss and glucose reduction during an oral glucose tolerance test (OGTT) following SC and intravenous (IV) administration in diet-induced obese (DIO) and leptin-deficient obese (ob/ob) mice, respectively. Our data show that the intact N-terminus of FGF21 in PF-05231023 appears to be sufficient to drive glucose lowering during OGTT and sustain BW loss in DIOs. Further, PK/PD modeling suggests that while the intact FGF21 C-terminus is not strictly required for glucose lowering during OGTT in ob/ob mice or for BW reduction in DIO mice, the higher potency conferred by intact C-terminus contributes to a rapid initiation of pharmacodynamic effects immediately following dosing. These results provide additional insight into the strategy of developing stabilized versions of FGF21 analogs to harness the full spectrum of its metabolic benefits. Introduction Fibroblast growth factor 21 (FGF21) is a member of the FGF19 subfamily that was discovered to be a critical metabolic regulator for maintenance of Mouse monoclonal to KLHL11 glucose and lipid homeostasis [1], thus emerging as a promising novel class of therapeutic for complex metabolic diseases such as Verinurad type 2 diabetes (T2D) and obesity [2,3]. The beneficial effects of native FGF21 and FGF21 analogues in normalizing glucose and lipid homeostasis have been demonstrated in a variety of preclinical metabolic disease models, including DIO mice, ob/ob mice, db/db mice, diabetic NHP and obese NHP [4C8]. Consistent with the metabolic benefits observed upon pharmacological administration of FGF21 in preclinical species, a recent clinical trial demonstrated robust effects of a stabilized FGF21 analog, LY2405319, in reducing hyperlipidemia and promoting body weight loss in obese T2D subjects [2]. FGF21-mediated biological effects are believed to be mediated through formation of an FGF21/ beta-klotho (KLB)/FGF receptor 1c FGFR1c complex and subsequent signaling [9,5,10]. Verinurad Native FGF21 is composed of 181 amino acids with a -trefoil core structure that is conserved in other FGF family proteins and free C- and N-termini that are unique to FGF21 [11,12,10]. The C-terminus of FGF21 is susceptible to proteolytic cleavage and the resultant metabolite is ~200 fold less potent in vitro [13C15]. Moreover, the in vivo half-life of intact native FGF21 is less than 2 hr across multiple species and therefore not ideal for development as a therapeutic for chronic metabolic diseases, such as T2D and obesity. As a result, a number of long-acting FGF21 analogs, including protease stabilized protein [7], Fc-fusion [16], PEG-conjugate [17], and antibodies have been generated and tested in a variety of preclinical species to harness the metabolic benefits of the molecule [18]. PF-05231023 is a long-acting FGF21 analog developed by conjugating two molecules of modified FGF21 [dHis/Ala129Cys] to an antibody scaffold, CovX-2000 [6]. The pharmacokinetics (PK) of PF-05231023 molecule were characterized using an ELISA that measures exposure of drug-related molecules containing the mid-region of FGF21 molecule attached to CovX-2000. Data from ELISA showed PF-05231023 had a prolonged in vivo half-life compared to native FGF21. However,.