Monitoring the cytosolic entry of cell-penetrating peptides using a pH-sensitive fluorophore

Abstract

We report a simple, effective method to assess the cytosolic delivery efficiency and kinetics of cell-penetrating peptides using a pH-sensitive fluorescent probe, naphthofluorescein.

Enhancing Antitumor Immune Responses by Optimized Combinations of Cell-penetrating Peptide-based Vaccines and Adjuvants

“…23,24 Further, in vivo studies would be therefore necessary to determine whether the low level of CD8 T cell immune responses induced by Z15 CPP was due to inefficient escape from the endosome to the cytosol. 27 Altogether, the results point out that the level of efficacy of a CPPbased vaccine cannot be uniquely related to its in vitro capacities. Whereas Z18 is a potent ZEBRA CPP truncation when conjugated to CD4 and CD8 epitopes of OVA, it was less potent to elicit gp100-specific CD8 T cells when conjugated to this epitope and the OVA CD4 epitope (Figure 8a,b).…”

“…To extend the previous results, OVACD8 epitope was replaced by the TAA gp100 [25][26][27][28][29][30][31][32][33] CD8 epitope in the previous constructs (see Supplementary Table S5) to evaluate the TAA-specific immune response. These constructs were tested with Hiltonol (Figure 8a) or MPLA adjuvant (Figure 8b).…”

“…with 1 × 10 5 B16 cells expressing gp100 and OVA antigens. Vaccinations were made with ZEBRA CPP variant vaccines expressing gp100 [25][26][27][28][29][30][31][32][33] CD8 epitope and the OVA323-339 CD4 epitope (Figure 8c). Z14 CPP was the most efficacious variant, supporting previous immune response data (Figure 8a).…”

Cell-permeable bicyclic peptidyl inhibitors against T-cell protein tyrosine phosphatase from a combinatorial library

“…Because NF has a pKa value of 7.8 and is non-fluorescent inside the acidic endosomes and lysosomes (pH ≤6), it has been used to effectively quantitate the cytosolic entry of CPPs. 10,15 The flow cytometry data indicate that peptides 25 and 33 enter the cytosol and nucleus of HeLa cells with 35% and 180% efficiencies, respectively (an efficiency of 100% is defined as equal peptide concentration in the extracellular growth medium and the cytosol) (Fig. 3C).…”

Intracellular Delivery of Proteins By Nanocarriers

“…As a result, the amount of protein reaching the cytosol is low, with concomitantly low efficacy of the delivered protein [8]. Although, progress has been made in increasing the efficiency of endosomal escape of delivered proteins [9], these methods are still generally of rather low efficiency [10].…”