Alice Bakker

Multivalent avimer proteins evolved by exon shuffling of a family of human receptor domains

Nat. Biotechnol. 23, 1189–1190 (2005) In the print version of this article and the version originally published online, a company name was misspelled. On page 1190, Box 1, paragraph 1, line 3, “Macrogene” should have been “Macrogen.” This correction is appended to the PDF version.

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. In a study conducted in obese male cynomolgus monkeys, animals treated with an FGFR1c/β-Klotho bispecific Avimer showed improved metabolic parameters and reduced body weight comparable to the effects seen with FGF21. These results not only demonstrate the essential roles of FGFR1c and β-Klotho in mediating the metabolic effects of FGF21, they also describe a first bispecific activator of this unique receptor complex and provide validation for a novel therapeutic approach to target this potentially important pathway for treating diabetes and obesity.

A NOVEL PROTEIN THERAPEUTIC JOINT RETENTION STRATEGY BASED ON COLLAGEN-BINDING AVIMERS

Designing drugs to treat diseases associated with articular joints, particularly those targeting chondrocytes, is challenging due to unique local environmental constraints including the avascular nature of cartilage, the absence of a closed joint compartment, and a highly cross‐linked extracellular matrix. In an effort to address these challenges, we developed a novel strategy to prolong residence time of intra‐articularly administered protein therapeutics. Avimer domains are naturally found in membrane polypeptides and mediate diverse protein–protein interactions. Screening of a phage Avimer domain library led to identification of several low affinity type II collagen‐binding Avimers. Following several rounds of mutagenesis and reselection, these initial hits were transformed to high affinity, selective type II collagen‐binding Avimers. One such Avimer (M26) persisted in rat knees for at least 1 month following intra‐articular administration. Fusion of this Avimer to a candidate therapeutic payload, IL‐1Ra, yielded a protein construct which simultaneously bound to type II collagen and to IL‐1 receptor. In vitro, IL‐1Ra_M26 bound selectively to cartilage explants and remained associated even after extensive washing. Binding appeared to occur preferentially to pericellular regions surrounding chondrocytes. An acute intra‐articular IL‐1‐induced IL‐6 challenge rat model was employed to assess in vivo pharmacodynamics. Whereas both IL‐1Ra_M26 and native IL‐1Ra inhibited IL‐6 output when co‐administered with the IL‐1 challenge, only IL‐1Ra_M26 inhibited when administered 1 week prior to IL‐1 challenge. Collagen‐binding Avimers thus represent a promising strategy for enhancing cartilage residence time of protein therapeutics.

Abstract B233: c‐Met antagonist Avimer™ polypeptide that exhibits potent in vivo and in vitro inhibitor activity

c‐Met is a receptor tyrosine kinase (RTK) that is activated upon binding to its only known ligand, hepatocyte growth factor (HGF). Although essential in development, aberrant c‐Met signaling can lead to oncogenesis by promoting proliferation, survival, metastasis, and angiogenesis. Deregulated expression/activation of cMet is found in many neoplasms, and the linkage to disease severity and poor prognosis make this RTK an attractive candidate for targeted anticancer therapy. Challenges of inhibiting c‐Met/HGF signaling with a classic antibody therapeutic approach have included: (1) the inhibition of receptor activation required more than one antibody, and (2) bivalent antibodies caused receptor activation. Using a novel non‐antibody protein technology platform, we have identified potent polypeptide inhibitors of HGF‐mediated c‐Met activation. This non‐antibody class of proteins, termed Avimer (avidity multimer) polypeptides, are based on naturally occurring domain (e.g., A‐domain) containing proteins and can be generated using a phage display library and expressed in E. coli . c‐Met antagonist Avimer polypeptides potently inhibit HGF‐mediated receptor activation, migration, and proliferation in in vitro assays. Improvement in pharmacokinetic profile gained through attaching polyethylene glycol molecules to avimers enabled in vivo studies. In multiple preclinical tumor models (U87MG glioblastoma, KP‐4 pancreatic, U118 glioblastoma), Avimer‐treated animals exhibited reduced cMet activation in xenografts and significantly decreased tumor growth. These data indicate that the c‐Met antagonist Avimer polypeptides are potent c‐Met inhibitors and offer an alternative to small molecules or antibodies in targeting cMet/HGF pathway for anticancer therapy. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B233.