title: Optimization and in Vivo Validation of Peptide Vectors Targeting the LDL Receptor.
abstract: Active targeting and delivery to pathophysiological organs of interest is of paramount importance to increase specific accumulation of therapeutic drugs or imaging agents while avoiding systemic side effects. We recently developed a family of new peptide ligands of the human and rodent LDL receptor (LDLR), an attractive cell-surface receptor with high uptake activity and local enrichment in several normal or pathological tissues (Malcor et al., J. Med. Chem. 2012, 55 (5), 2227). Initial chemical optimization of the 15-mer, all natural amino acid compound 1/VH411 (DSGL[CMPRLRGC]cDPR) and structure-activity relationship (SAR) investigation led to the cyclic 8 amino acid analogue compound 22/VH445 ([cMPRLRGC]c) which specifically binds hLDLR with a KDof 76 nM and has an in vitro blood half-life of ∼3 h. Further introduction of non-natural amino acids led to the identification of compound 60/VH4106 ([(d)-"Pen"M"Thz"RLRGC]c), which showed the highest KDvalue of 9 nM. However, this latter analogue displayed the lowest in vitro blood half-life (∼1.9 h). In the present study, we designed a new set of peptide analogues, namely, VH4127 to VH4131, with further improved biological properties. Detailed analysis of the hLDLR-binding kinetics of previous and new analogues showed that the latter all displayed very high on-rates, in the 106s-1.M-1range, and off-rates varying from the low 10-2s-1to the 10-1s-1range. Furthermore, all these new analogues showed increased blood half-lives in vitro, reaching ∼7 and 10 h for VH4129 and VH4131, respectively. Interestingly, we demonstrate in cell-based assays using both VH445 and the most balanced optimized analogue VH4127 ([cM"Thz"RLRG"Pen"]c), showing a KDof 18 nM and a blood half-life of ∼4.3 h, that its higher on-rate correlated with a significant increase in both the extent of cell-surface binding to hLDLR and the endocytosis potential. Finally, intravenous injection of tritium-radiolabeled3H-VH4127 in wild-type or ldlr -/- mice confirmed their active LDLR targeting in vivo. Overall, this study extends our previous work toward a diversified portfolio of LDLR-targeted peptide vectors with validated LDLR-targeting potential in vivo.
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Jacquot
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Pascaline
authors.initials: P
authors.lastname: Lécorché
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Jean-Daniel
authors.initials: JD
authors.lastname: Malcor
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.|CEA-Saclay, Service d'Ingénierie Moléculaire de Protéines (SIMOPRO), Labex LERMIT, CEA-DSV-iBiTecS , 91191 Gif/Yvette Cedex, France.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Laurencin
authors.affiliation: Variabilité de réponse aux psychotropes, INSERM U1144 , Paris 75006, France.|Faculté de Pharmacie, Université Paris Descartes, INSERM UMR S-1144 , Paris 75006, France.
authors.forename: Maria
authors.initials: M
authors.lastname: Smirnova
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.|Aix Marseille Univ, CNRS, NICN , Marseille, France.
authors.forename: Karine
authors.initials: K
authors.lastname: Varini
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Cédric
authors.initials: C
authors.lastname: Malicet
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Fanny
authors.initials: F
authors.lastname: Gassiot
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Karima
authors.initials: K
authors.lastname: Abouzid
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Aude
authors.initials: A
authors.lastname: Faucon
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Marion
authors.initials: M
authors.lastname: David
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.|Aix Marseille Univ, CNRS, NICN , Marseille, France.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Gaudin
authors.affiliation: VECT-HORUS SAS, Faculté de Médecine secteur Nord , 51 Boulevard Pierre Dramard, CS80011, 13344 Marseille Cedex 15, France.
authors.forename: Maxime
authors.initials: M
authors.lastname: Masse
authors.affiliation: Aix Marseille Univ, CNRS, CRN2M , Marseille, France.
authors.forename: Géraldine
authors.initials: G
authors.lastname: Ferracci
authors.affiliation: CEA-Saclay, Service d'Ingénierie Moléculaire de Protéines (SIMOPRO), Labex LERMIT, CEA-DSV-iBiTecS , 91191 Gif/Yvette Cedex, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Dive
authors.affiliation: Variabilité de réponse aux psychotropes, INSERM U1144 , Paris 75006, France.|Faculté de Pharmacie, Université Paris Descartes, INSERM UMR S-1144 , Paris 75006, France.
authors.forename: Salvatore
authors.initials: S
authors.lastname: Cisternino
authors.affiliation: Aix Marseille Univ, CNRS, NICN , Marseille, France.
authors.forename: Michel
authors.initials: M
authors.lastname: Khrestchatisky
mesh_terms: D000818:Animals; D016466:CHO Cells; D003412:Cricetulus; D016503:Drug Delivery Systems; D004705:Endocytosis; D005822:Genetic Vectors / Q000592:standards*; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010446:Peptide Fragments / Q000493:pharmacokinetics / Q000494:pharmacology*; D011973:Receptors, LDL / Q000037:antagonists & inhibitors* / Q000502:physiology; D013329:Structure-Activity Relationship; D014018:Tissue Distribution
chemical_list: D010446:Peptide Fragments; D011973:Receptors, LDL
keywords: LDLR knockout; LDLR targeting; medicinal chemistry based optimization; peptide vector; tissue distribution