The ANGPTL3-ANGPTL8 bispecific antibody attenuates adriamycin-induced chronic nephritis-like renal injury and fibrosis by suppressing the integrin αvβ3-STAT3 signaling axis

Chronic glomerulonephritis (CGN) is characterized by persistent renal inflammation and fibrosis, but its pathological spectrum is heterogeneous. Adriamycin-induced nephropathy is a commonly used podocyte injury-associated model that develops albuminuria, renal injury, and secondary inflammatory and fibrotic changes. Angiopoietin-like protein 3 (ANGPTL3) and angiopoietin-like protein 8 (ANGPTL8) are implicated in renal pathology, but their coordinated involvement and therapeutic targeting remain incompletely defined. This study aimed to construct an ANGPTL3-ANGPTL8 bispecific antibody (A3-A8) and evaluate its protective effects in an adriamycin-induced chronic nephritis-like model through the integrin αvβ3-STAT3 signaling axis. A rat adriamycin-induced nephropathy model was established as an experimental model of chronic nephritis-like renal injury, and glomerular RNA sequencing (RNA-seq) was performed to identify differentially expressed genes. The A3-A8 bispecific antibody was expressed in E. coli, purified, and characterized for thermal stability and binding affinity by surface plasmon resonance. In vitro s tudies used lipopolysaccharide (LPS)-treated human renal fibroblast cells with Cell Counting Kit-8 (CCK-8), flow cytometry, enzyme-linked immunosorbent assay (ELISA), immunofluorescence, and Western blot analyses. shRNA-mediated knockdown and overexpression were used for mechanistic validation. In vivo efficacy was assessed in model rats treated with A3-A8 with or without osteopontin (OPN), and additional renal tissue signaling, podocyte injury, albuminuria, and cilengitide intervention analyses were performed. ANGPTL3 and ANGPTL8 were markedly upregulated in the adriamycin-induced renal injury model. The bispecific antibody showed high thermal stability (Tm1 = 66.6 °C) and nanomolar-range binding affinities to both rat and human ANGPTL3 and ANGPTL8. It dose-dependently reduced apoptosis, inflammation (IL-1β, IL-6, TNF-α), and fibrosis markers (collagen I, fibronectin), while increasing cell viability. A3-A8 suppressed integrin αvβ3 and p-STAT3; αvβ3 overexpression reversed its effects, which were blocked by STAT3 knockdown. In vivo, A3-A8 attenuated renal fibrosis, inflammatory infiltration, albuminuria, podocyte injury, and renal dysfunction. Renal tissue Western blot (WB)/immunohistochemistry (IHC) and cilengitide intervention further supported involvement of the αvβ3-STAT3 axis. The A3-A8 bispecific antibody alleviates adriamycin-induced chronic nephritis-like renal injury and fibrosis, at least partly by inhibiting the integrin αvβ3-STAT3 axis, and represents a potential bispecific therapeutic candidate.