FGF21 trafficking in intact human cells revealed by cryo-electron tomography with gold nanoparticles.

The fibroblast growth factor FGF21 was labeled with molecularly defined gold nanoparticles (AuNPs), applied to human adipocytes, and imaged by cryo-electron tomography (cryo-ET). Most AuNPs were in pairs about 80 Å apart, on the outer cell surface. Pairs of AuNPs were also abundant inside the cells in clathrin-coated vesicles and endosomes.

Agonistic β-Klotho antibody mimics fibroblast growth factor 21 (FGF21) functions.

Fibroblast growth factor 21 (FGF21), an endocrine hormone in the FGF family, plays a critical role in regulating metabolic homeostasis and has emerged as a therapeutic target for metabolic diseases, including Type 2 diabetes mellitus. FGF21 functions through a receptor complex that consists of an FGF receptor (FGFR) and a co-receptor β-Klotho. Here, we identify and biochemically and structurally characterize 39F7, a high-affinity agonistic monoclonal antibody (mAb) against β-Klotho that mimics FGF21 function.

A systematic dissection of sequence elements determining β-Klotho and FGF interaction and signaling.

Endocrine fibroblast growth factors (FGFs) require Klotho transmembrane proteins as necessary co-receptors to activate FGF receptor (FGFR) signaling. In particular, FGF19 and FGF21 function through β-Klotho to regulate glucose and lipid metabolism. Recent research has focused on elucidating how these two FGFs interact with β-Klotho and FGFRs to activate downstream signaling.

Impact of antibody subclass and disulfide isoform differences on the biological activity of CD200R and βklotho agonist antibodies.

Agonism of cell surface receptors by monoclonal antibodies is dependent not only on its ability to bind the target, but also to deliver a biological signal through receptors to the cell. Immunoglobulin G2 antibodies (IgG2s) are made up of a mixture of distinct isoforms (IgG2-A, -B and A/B), which differ by the disulfide connectivity at the hinge region. When evaluating panels of agonistic antibodies against CD200 receptor (CD200R) or βklotho receptor (βklotho), we noticed striking activity differences of IgG1 or IgG2 antibodies with the same variable domains.

A Novel Fc-FGF21 With Improved Resistance to Proteolysis, Increased Affinity Toward β-Klotho, and Enhanced Efficacy in Mice and Cynomolgus Monkeys.

Fibroblast growth factor (FGF) 21 is a natural hormone that modulates glucose, lipid, and energy metabolism. Previously, we engineered an Fc fusion FGF21 variant with two mutations, Fc-FGF21(RG), to extend the half-life and reduce aggregation and in vivo degradation of FGF21. We now describe a new variant developed to reduce the extreme C-terminal degradation and improve the binding affinity to β-Klotho.

Chronic Over-expression of Fibroblast Growth Factor 21 Increases Bile Acid Biosynthesis by Opposing FGF15/19 Action.

Pharmacological doses of fibroblast growth factor (FGF) 21 effectively normalize glucose, lipid and energy homeostasis in multiple animal models with many benefits translating to obese humans with type 2 diabetes. However, a role for FGF21 in the regulation of bile acid metabolism has not been reported. Herein, we demonstrate AAV-mediated FGF21 overexpression in mice increases liver expression of the key bile acid producing enzyme, Cyp7a1, resulting in an increased bile acid pool.

Fibroblast growth factor receptor 4 (FGFR4) deficiency improves insulin resistance and glucose metabolism under diet-induced obesity conditions.

The role of fibroblast growth factor receptor 4 (FGFR4) in regulating bile acid synthesis has been well defined; however, its reported role on glucose and energy metabolism remains unresolved. Here, we show that FGFR4 deficiency in mice leads to improvement in glucose metabolism, insulin sensitivity, and reduction in body weight under high fat conditions. Mechanism of action studies in FGFR4-deficient mice suggest that the effects are mediated in part by increased plasma levels of adiponectin and the endocrine FGF factors FGF21 and FGF15, the latter of which increase in response to an elevated bile acid pool.

FGF21 can be mimicked in vitro and in vivo by a novel anti-FGFR1c/β-Klotho bispecific protein.

The endocrine hormone FGF21 has attracted considerable interest as a potential therapeutic for treating diabetes and obesity. As an alternative to the native cytokine, we generated bispecific Avimer polypeptides that bind with high affinity and specificity to one of the receptor and coreceptor pairs used by FGF21, FGFR1c and β-Klotho. These Avimers exhibit FGF21-like activity in in vitro assays with potency greater than FGF21.

A novel approach to improve the function of FGF21.

Background And Objective: Fibroblast growth factor 21 (FGF21) has potent effects on normalizing glucose, lipid, and energy homeostasis, and represents an attractive novel therapy for type 2 diabetes mellitus and obesity. Approaches to improve the pharmacokinetic properties of FGF21, such as conjugation with polyethylene glycol, have been explored for therapeutic development. However, not only is there room for further pharmacokinetic improvements, additional re-engineering approaches to improve the potency and stability of FGF21 have not been reported.

Discovery of INT131: a selective PPARγ modulator that enhances insulin sensitivity.

PPARγ is a member of the nuclear hormone receptor family and plays a key role in the regulation of glucose homeostasis. This Letter describes the discovery of a novel chemical class of diarylsulfonamide partial agonists that act as selective PPARγ modulators (SPPARγMs) and display a unique pharmacological profile compared to the thiazolidinedione (TZD) class of PPARγ full agonists. Herein we report the initial discovery of partial agonist 4 and the structure-activity relationship studies that led to the selection of clinical compound INT131 (3), a potent PPARγ partial agonist that displays robust glucose-lowering activity in rodent models of diabetes while exhibiting a reduced side-effects profile compared to marketed TZDs.

Dual actions of fibroblast growth factor 19 on lipid metabolism.

Elevated triglyceride (TG) and cholesterol levels are risk factors for cardiovascular disease and are often associated with diabetes and metabolic syndrome. Recent reports suggest that fibroblast growth factor (FGF)19 and FGF21 can dramatically improve metabolic dysfunction, including hyperglycemia, hypertriglyceridemia, and hypercholesterolemia. Due to their similar receptor specificities and co-receptor requirements, FGF19 and FGF21 share many common properties and have been thought to be interchangeable in metabolic regulation.

Treating diabetes and obesity with an FGF21-mimetic antibody activating the βKlotho/FGFR1c receptor complex.

Fibroblast growth factor 21 (FGF21) is a distinctive member of the FGF family with potent beneficial effects on lipid, body weight, and glucose metabolism and has attracted considerable interest as a potential therapeutic for treating diabetes and obesity. As an alternative to native FGF21, we have developed a monoclonal antibody, mimAb1, that binds to βKlotho with high affinity and specifically activates signaling from the βKlotho/FGFR1c (FGF receptor 1c) receptor complex. In obese cynomolgus monkeys, injection of mimAb1 led to FGF21-like metabolic effects, including decreases in body weight, plasma insulin, triglycerides, and glucose during tolerance testing.

Characterization of a FGF19 variant with altered receptor specificity revealed a central role for FGFR1c in the regulation of glucose metabolism.

Diabetes and associated metabolic conditions have reached pandemic proportions worldwide, and there is a clear unmet medical need for new therapies that are both effective and safe. FGF19 and FGF21 are distinctive members of the FGF family that function as endocrine hormones. Both have potent effects on normalizing glucose, lipid, and energy homeostasis, and therefore, represent attractive potential next generation therapies for combating the growing epidemics of type 2 diabetes and obesity.

A unique FGF23 with the ability to activate FGFR signaling through both αKlotho and βKlotho.

Three fibroblast growth factor (FGF) molecules, FGF19, FGF21, and FGF23, form a unique subfamily that functions as endocrine hormones. FGF19 and FGF21 can regulate glucose, lipid, and energy metabolism, while FGF23 regulates phosphate homeostasis. The FGF receptors and co-receptors for these three FGF molecules have been identified, and domains important for receptor interaction and specificity determination are beginning to be elucidated.

The FGFR D3 domain determines receptor selectivity for fibroblast growth factor 21.

FGF21 is a member of a unique subfamily of fibroblast growth factors that function as endocrine hormones and regulate a variety of metabolic activities. Unlike paracrine FGFs, FGF21 does not bind heparin and requires βKlotho as a co-receptor to activate FGFR signaling. In the presence of βKlotho, FGF21 is able to activate FGFRs 1c, 2c and 3c but not FGFR4.

Mutational analysis of G-protein coupled receptor--FFA2.

FFA2 (GPR43) is a receptor for short-chain fatty acids (SCFAs), acetate, and propionate. FFA2 is predominantly expressed in islets, a subset of immune cells, adipocytes, and the gastrointestinal tract which suggest a possible role in inflammatory and metabolic conditions. We have previously described the identification and characterization of novel phenylacetamides as allosteric agonists of FFA2.

Allosteric rescuing of loss-of-function FFAR2 mutations.

FFAR2 (GPR43) is a receptor for short-chain fatty acids (SCFAs), acetate and propionate. In the current study, we investigate the molecular determinants contributing to receptor activation by endogenous ligands. Mutational analysis revealed several important residues located in transmembrane domains (TM) 3, 4, 5, 6, and 7 for acetate binding.

Separating mitogenic and metabolic activities of fibroblast growth factor 19 (FGF19).

FGF19 and FGF21 are distinctive members of the FGF family that function as endocrine hormones. Their potent effects on normalizing glucose, lipid, and energy homeostasis in disease models have made them an interesting focus of research for combating the growing epidemics of diabetes and obesity. Despite overlapping functions, FGF19 and FGF21 have many discrete effects, the most important being that FGF19 has both metabolic and proliferative effects, whereas FGF21 has only metabolic effects.

Generation of novel long-acting globular adiponectin molecules.

Adiponectin is an adipocyte-derived hormone that has been shown to play important roles in the regulation of glucose and energy homeostasis. It exists as homotrimers or complexes containing multiple homotrimer units in plasma. The recombinant adiponectin proteins have been difficult to produce, making it challenging for both research as well as potential therapeutic development.

FGF19-induced hepatocyte proliferation is mediated through FGFR4 activation.

FGF19 and FGF21, unique members of the fibroblast growth factor (FGF) family, are hormones that regulate glucose, lipid, and energy homeostasis. Increased hepatocyte proliferation and liver tumor formation have also been observed in FGF19 transgenic mice. Here, we report that, in contrast to FGF19, FGF21 does not induce hepatocyte proliferation in vivo.

The first synthetic agonists of FFA2: Discovery and SAR of phenylacetamides as allosteric modulators.

Free fatty acid receptor 2 (FFA2) is a G-protein coupled receptor for which only short-chain fatty acids (SCFAs) have been reported as endogenous ligands. We describe the discovery and optimization of phenylacetamides as allosteric agonists of FFA2. These novel ligands can suppress adipocyte lipolysis in vitro and reduce plasma FFA levels in vivo, suggesting that these allosteric modulators can serve as pharmacological tools for exploring the potential function of FFA2 in various disease conditions.

Inhibition of lipolysis may contribute to the acute regulation of plasma FFA and glucose by FGF21 in ob/ob mice.

FGF21 is a unique member of the fibroblast growth factors (FGFs) and a novel hormone that regulates glucose, lipid, and energy homeostasis. The beneficial effects of FGF21 reported thus far have mostly been from chronic treatments. In order to better understand the mechanism for FGF21 action, we evaluated the acute effects of FGF21 in vivo and in vitro.

Selective activation of FGFR4 by an FGF19 variant does not improve glucose metabolism in ob/ob mice.

FGF19 is a hormone that regulates bile acid and glucose homeostasis. Progress has been made in identifying cofactors for receptor activation. However, several functions of FGF19 have not yet been fully defined, including the actions of FGF19 on target tissues, its FGF receptor specificity, and the contributions of other cofactors, such as heparin.

INT131: a selective modulator of PPAR gamma.

The nuclear hormone receptor peroxisome proliferator-activated receptor gamma (PPAR gamma; NR1C3) plays a central role in adipogenesis and is the molecular target of the thiazolidinedione class of antidiabetic drugs. To overcome the well-known shortcomings of thiazolidinediones, we have identified INT131 (formerly T131 and AMG131) as a potent selective ligand for PPAR gamma that is structurally and pharmacologically distinct from glitazone agonists. In vitro biochemical and cell-based functional assays showed that INT131 mediates a distinct pattern of coregulator recruitment to PPAR gamma.

C-terminal tail of FGF19 determines its specificity toward Klotho co-receptors.

FGF19 subfamily proteins (FGF19, FGF21, and FGF23) are unique members of fibroblast growth factors (FGFs) that regulate energy, bile acid, glucose, lipid, phosphate, and vitamin D homeostasis in an endocrine fashion. Their activities require the presence of alpha or betaKlotho, two related single-pass transmembrane proteins, as co-receptors in relevant target tissues. We previously showed that FGF19 can bind to both alpha and betaKlotho, whereas FGF21 and FGF23 can bind only to either betaKlotho or alphaKlotho, respectively in vitro.