Highway to Cell: Selection of the Best Cell-Penetrating Peptide to Internalize the CFTR-Stabilizing iCAL36 Peptide

Abstract

Therapeutic peptides have regained interest as they can address unmet medical needs and can be an excellent complement to pharmaceutic small molecules and other macromolecular therapeutics. Over the past decades, correctors and potentiators of the cystic fibrosis transmembrane conductance regulator (CFTR), a chloride ion channel causing cystic fibrosis (CF) when mutated, were developed to reduce the symptoms of the patients. In this context, we have previously designed a CFTR-stabilizing iCAL36 peptide able to further increase the CFTR amount in epithelial cells, thereby resulting in a higher CFTR activity. In the present study, optimization of the peptidyl inhibitor was performed by coupling five different cell-penetrating peptides (CPP), which are Tat, dTat, TatRI (retro-inverso), MPG, and Penetratin. Screening of the internalization properties of these CPP-iCAL36 peptides under different conditions (with or without serum or endocytosis inhibitors, etc.) was performed to select TatRI as the optimal CPP for iCAL36 delivery. More importantly, using this TatRI-iCAL36 peptide, we were able to reveal for the first time an additive increase in the CFTR amount in the presence of VX-445/VX-809 compared to VX-445/VX-809 treatment alone. This finding is a significant contribution to the development of CFTR-stabilizing peptides in addition to currently used treatments (small-molecule correctors or potentiators) for CF patients.

Cell Penetrating Peptides: Classification, Mechanisms, Methods of Study, and Applications

“…Synthetic designed S5-S5 stapled peptides, alpha helical [127] TatRI qpprrrqrrkkrg Designed to deliver a iCal36 CFTR [a] stabilizing peptide Retro-inverso [128] diLR10 LCKLLKKLCK Synthetic designed Alpha helical, self-assembling in a dimeric form [129] [a] Cystic fibrosis transmembrane conductance regulator.…”

A Proteomic Survey of the Cystic Fibrosis Transmembrane Conductance Regulator Surfaceome

“…The transgenic expression of GRASP in ∆F508-CFTR expressing mice has been shown to restore channel function with no observable toxicity [24]. The previously mentioned GOPC protein contains two coiled-coil domains and one PDZ domain that are known to interact with the C-terminus of CFTR [66,67]. The interaction between GOPC:CFTR reduces surface CFTR through its endocytic recycling by targeting it for lysosomal degradation, and inhibiting this interaction helps stabilize CFTR at the PM [67].…”

“…The previously mentioned GOPC protein contains two coiled-coil domains and one PDZ domain that are known to interact with the C-terminus of CFTR [66,67]. The interaction between GOPC:CFTR reduces surface CFTR through its endocytic recycling by targeting it for lysosomal degradation, and inhibiting this interaction helps stabilize CFTR at the PM [67]. Inhibiting GOPC was also reported to have an additive increase in the surface quantity of CFTR in the presence of front-line small molecular correctors, VX445/VX-809 [67].…”

“…The interaction between GOPC:CFTR reduces surface CFTR through its endocytic recycling by targeting it for lysosomal degradation, and inhibiting this interaction helps stabilize CFTR at the PM [67]. Inhibiting GOPC was also reported to have an additive increase in the surface quantity of CFTR in the presence of front-line small molecular correctors, VX445/VX-809 [67]. The Scribble cell polarity module, comprising Scribbled (Scrib), Discs-large (Dlg), and Lethal-2-giant larvae (Lgl), has a tumor suppressive role in mammalian epithelial cancers [68].…”

Topoisomeric Membrane-Active Peptides: A Review of the Last Two Decades

“…Wender et al studied a shortened version, Tat [49][50][51][52][53][54][55][56][57], as well as the corresponding e and re versions and found that these topoisomers were more effective than the canonic L-version in terms of entering Jurkat cells [191]. Similarly, Seisel et al investigated the e and re versions of the Tat [48][49][50][51][52][53][54][55][56][57][58][59][60] sequence using iCal36, a peptide intended for patients with cystic fibrosis, as cargo, with the re topoisomer found to be most appropriate [192].…”