The immunophilin protein FKBPL and its peptide derivatives are novel regulators of vascular integrity and inflammation via NF-κB signaling

Abstract

AbstractA breakdown in vascular integrity and excessive inflammation are hallmarks of serious pathological conditions including sepsis, acute respiratory distress syndrome (ARDs) and most recently, severe COVID-19. FK506 – binding protein like (FKBPL) is a member of the immunophilin protein superfamily with potent anti-tumor activity through inhibition of angiogenesis and cancer stemness. An FKBPL-based 23mer peptide, ALM201, displayed a good safety and pharmacokinetic profile in a Phase 1a oncology clinical trial and was subsequently designated orphan drug status by the FDA in ovarian cancer. Here we describe a novel role for FKBPL and its peptides in regulating vascular integrity and cytokine production though modulating NF-κB signaling. FKBPL knockdown promoted endothelial cell barrier permeability, which was further exacerbated upon stimulation with lipopolysaccharide (LPS) and accompanied by increased expression of TNF mRNA and phosphorylation of p65(RelA). Whilst treatment with the FKBPL based pre-clinical peptide, AD-01, increased VE-cadherin endothelial tight junctions following LPS stimulation. Bone marrow derived macrophages (BMDM) from FKBPL haploinsufficient mice (Fkbpl+/−) also demonstrated increased phosphorylation of p65(RelA) in response to LPS stimulation compared to wild-type mice. Furthermore, treatment with AD-01 inhibited p65(RelA) phosphorylation following LPS stimulation resulting in reduced NF-κB target gene expression and proinflammatory cytokine production. In an in vivo LPS survival model, Fkbpl+/− mice have reduced survival compared to wild-type mice. Moreover, treatment of wild-type mice with the clinical FKBPL-based peptide, ALM201, following LPS injection resulted in a 100% survival rate in mice at experimental endpoint, as well as an abrogation of production of pro-inflammatory cytokines, TNF and IL-6, in peritoneal lavage washings. Analysis of human genetic biobanks found an association between common genetic variants associated with FKBPL and traits associated with inflammatory disorders such as psoriasis, rheumatoid arthritis and high lymphocyte count. In summary, for the first time, we describe a novel role for FKBPL as a regulator of inflammation and vascular integrity through modulating NF-κB signaling and FKBPL based therapies demonstrate potent anti-inflammatory activity.