Pharmacokinetic Engineering of OX40-Blocking Anticalin Proteins Using Monomeric Plasma Half-Life Extension Domains

Frontiers in Pharmacology

Martin Siegemund¹  Karoline Utschick²  Christina Grasmüller²  Gunther Galler²  Christine Rothe²  Andrea Allersdorfer²  Prajakta Oak²  Alexandra Winter²  Eva-Maria Hansbauer²  Josef Prassler²  Jan-Peter Mayer²  Benjamin Weiche²  Roland E. Kontermann³ 

[1] Institute of Cell Biology and Immunology, University of Stuttgart, Stuttgart, Germany;Pieris Pharmaceuticals GmbH, Hallbergmoos, Germany;Stuttgart Research Center Systems Biology, University of Stuttgart, Stuttgart, Germany;

关键词: Anticalin protein;    scaffold protein;    half-life extension;    albumin;    IgG;    binding domain;   

DOI  :  10.3389/fphar.2021.759337

来源: DOAJ

【 摘 要 】

Anticalin® proteins have been proven as versatile clinical stage biotherapeutics. Due to their small size (∼20 kDa), they harbor a short intrinsic plasma half-life which can be extended, e.g., by fusion with IgG or Fc. However, for antagonism of co-immunostimulatory Tumor Necrosis Factor Receptor Superfamily (TNFRSF) members in therapy of autoimmune and inflammatory diseases, a monovalent, pharmacokinetically optimized Anticalin protein format that avoids receptor clustering and therefore potential activation is favored. We investigated the suitability of an affinity-improved streptococcal Albumin-Binding Domain (ABD) and the engineered Fab-selective Immunoglobulin-Binding Domain (IgBD) SpGC3Fab for plasma Half-Life Extension (HLE) of an OX40-specific Anticalin and bispecific Duocalin proteins, neutralizing OX40 and a second co-immunostimulatory TNFRSF member. The higher affinity of ABD fusion proteins to human serum albumin (HSA) and Mouse Serum Albumin (MSA), with a 4 to 5-order of magnitude lower KD compared with the binding affinity of IgBD fusions to human/mouse IgG, translated into longer terminal plasma half-lives (t1/2). Hence, the anti-OX40 Anticalin-ABD protein reached t1/2 values of ∼40 h in wild-type mice and 110 h in hSA/hFcRn double humanized mice, in contrast to ∼7 h observed for anti-OX40 Anticalin-IgBD in wild-type mice. The pharmacokinetics of an anti-OX40 Anticalin-Fc fusion protein was the longest in both models (t1/2 of 130 h and 146 h, respectively). Protein formats composed of two ABDs or IgBDs instead of one single HLE domain clearly showed longer presence in the circulation. Importantly, Anticalin-ABD and -IgBD fusions showed OX40 receptor binding and functional competition with OX40L-induced cellular reactivity in the presence of albumin or IgG, respectively. Our results suggest that fusion to ABD or IgBD can be a versatile platform to tune the plasma half-life of Anticalin proteins in response to therapeutic needs.

【 授权许可】

Unknown