Second Generation I-Body AD-214 Attenuates Unilateral Ureteral Obstruction (UUO)-Induced Kidney Fibrosis Through Inhibiting Leukocyte Infiltration and Macrophage Migration

Abstract

Kidney fibrosis is the common pathological pathway in progressive chronic kidney disease (CKD), and current treatments are largely ineffective. The C-X-C chemokine receptor 4 (CXCR4) is crucial to fibrosis development. By using neural cell adhesion molecules as scaffolds with binding loops that mimic the shape of shark antibodies, fully humanized single-domain i-bodies have been developed. The first-generation i-body, AD-114, demonstrated antifibrotic effects in a mouse model of folic acid (FA)-induced renal fibrosis. The second-generation i-body, AD-214, is an Fc-fusion protein with an extended half-life, enhanced activity, and a mutated Fc domain to prevent immune activation. To investigate the renoprotective mechanisms of AD-214, RPTEC/TERT1 cells (a human proximal tubular cell line) were incubated with TGF-b1 with/without AD-214 and the supernatant was collected to measure collagen levels by Western blot. Mice with unilateral ureteral obstruction (UUO) received AD-214 intraperitoneally (i.p.) every two days for 14 days. Kidney fibrosis markers and kidney function were then analyzed. AD-214 suppressed TGF-b1-induced collagen overexpression in RPTEC/TERT1 cells. In UUO mice, AD-214 reduced extracellular matrix (ECM) deposition, restored kidney function, and limited leukocyte infiltration. In a scratch assay, AD-214 also inhibited macrophage migration. To conclude, i-body AD-214 attenuates UUO-induced kidney fibrosis by inhibiting leukocyte infiltration and macrophage migration.