Involvement of the low‐density lipoprotein receptor‐related protein in the transcytosis of the brain delivery vector Angiopep‐2

Abstract

The blood–brain barrier (BBB) restricts the entry of proteins as well as potential drugs to cerebral tissues. We previously reported that a family of Kunitz domain‐derived peptides called Angiopeps can be used as a drug delivery system for the brain. Here, we further characterize the transcytosis ability of these peptides using an in vitro model of the BBB and in situ brain perfusion. These peptides, and in particular Angiopep‐2, exhibited higher transcytosis capacity and parenchymal accumulation than do transferrin, lactoferrin, and avidin. Angiopep‐2 transport and accumulation in brain endothelial cells were unaffected by the P‐glycoprotein inhibitor, cyclosporin A, indicating that this peptide is not a substrate for the efflux pump P‐glycoprotein. However, competition studies show that activated α2‐macroglobulin, a specific ligand for the low‐density lipoprotein receptor‐related protein‐1 (LRP1) and Angiopep‐2 can share the same receptor. In addition, LRP1 was detected in glioblastomas and brain metastases from lung and skin cancers. Fluorescent microscopy also revealed that Alexa488‐Angiopep‐2 co‐localized with LRP1 in brain endothelial cell monolayers. Overall, these results suggest that Angiopep‐2 transport across the BBB is, in part, mediated by LRP1.

ANG4043, a Novel Brain-Penetrant Peptide–mAb Conjugate, Is Efficacious against HER2-Positive Intracranial Tumors in Mice

“…By conjugating An2 to the anti-HER2 mAb, we have created a bifunctional molecule that interacts with LRP1 as well as HER2. In previous studies, we reported colocalization between Alexa488-An2 and LRP1 in an in vitro bovine brain endothelial cell BBB model (23). In the current study, we demonstrate that ANG4043 uptake by HBMEC is pronounced and is attenuated in the presence of the LRP1 ligand, RAP.…”

“…LRP1 is a member of the LDL receptor family that is highly expressed on BBB capillary endothelial cells and has been shown to transport a variety of ligands, including lactoferrin, ApoE, and b-amyloid peptide, across brain capillary endothelial cells (20)(21)(22). We have previously described a 19-amino acid peptide, derived from the Kunitz domain of multiple LRP1 ligands, which binds LRP1 and efficiently penetrates the BBB by receptor-mediated transcytosis (23,24). Conjugation of this peptide, known as Angiopep-2 (An2), to small-molecule antineoplastic drugs like paclitaxel, doxorubicin, and etoposide, dramatically augments their uptake through the BBB following systemic administration (25,26).…”

Identification of blood protein biomarkers that aid in the clinical assessment of patients with malignant glioma

“…A2M has been found to be synthesized and secreted by human glioma cells and up-regulated in migrating human glioma cells compared with non-migrating glioma cells (21). The A2M receptor, LRP, is overexpressed in human malignant astrocytomas, especially in GBM (22), and A2M can cross the blood-brain barrier in an LRP-dependent manner (23). CA 19-9 is known to be a circulating protein biomarker for gastrointestinal cancers (24).…”

Shuttle–Cargo Fusion Molecules of Transport Peptides and the hD2/3Receptor Antagonist Fallypride: A Feasible Approach To Preserve Ligand–Receptor Binding?

“…The third subset of fusion molecules (III) was based on the AngioPep-2 peptide that enters the brain by specific receptor-mediated transcytosis utilizing the low-density lipoprotein related receptor LRP1. 17,44,45 Because it has been shown that a derivatization in position Lys 10 of the peptide is well-tolerated in terms of AngioPep's transport capacity, 16,18 the chelator DOTA was introduced in this position in all synthesized derivatives. For this purpose, the amino acid building block Fmoc−Lys(tris-tBu-DOTA)−OH (23) was synthesized and implemented into routine Fmoc-SPPS, giving improved yields compared to standard peptide scaffold synthesis, subsequent side-chain deprotection, and chelator functionalization.…”