Abstract
Therapeutics delivery into cells has been hurdled due to the barrier of cytoplasmic membrane. Although cell penetrating peptide (CPP) can potentially serve as an intracellular drug delivery vehicle, the application of CPP-based delivery is limited because the unsatisfactory delivery efficiency of CPP conjugated potent cargos is challenging their applications in present. Thus, the development of strategies for enhancing the penetrating efficiency of CPP would therefore urgent need to be explored to increase the scope of potential applications. We report here the effects of glucose, sucrose and manntiol (abbreviated as GSM) combination facilitating the penetration efficiency of CPP peptide alone or CPP-GFP (green fluorescence protein) conjugation in cultured cell lines or primary cells. Moreover, osmoprotectants glycerol and glycine supplementation help cells cope with the stress from GSM combination. Thus, our present study suggests that GSM combination in the presence of osmoprotectant can work as a new strategy for CPP penetration enhancement.
Highlights
Macromolecules including nucleic acids and proteins have demonstrated great value as research tools and achieved widespread success as human therapeutics among the fastest growing classes of drugs [1, 2]
A series of previous studies suggested that the suitable concentration of agents can promote the endosome-entrapped material release [15,16,17], in-vitro-cultured cells incubated with sucrose can serve as an alternative strategy to enhance the drug delivery
To further examine the enhancement effect of sucrose on the penetration of TAT, we detected the intracellular distribution of TAT using fluorescence microscope, as shown in Figure 1A, TAT-FITC was well-distributed in the cytosol of HeLa cells incubated with sucrose (500 mM), it seems like that enhancement effect of chloroquine with different concentration is limited (Supplementary Figure S1), and we detected intracellular distribution of TAT-FITC in different cells (Caski, A549, HepG2 and Siha) incubated with sucrose (500 mM) at 37°C for 1 h, as well as in Siha cells at 4°C (Figure 1B)
Summary
Macromolecules including nucleic acids (e.g. mRNA, siRNA, microRNA and lncRNA) and proteins (e.g. enzymes, cytokines and antibodies) have demonstrated great value as research tools and achieved widespread success as human therapeutics among the fastest growing classes of drugs [1, 2]. Many powerful and potentially therapeutic proteins are typically not able to spontaneously enter into mammalian cells owing to the selectively permeable barrier of cellular membrane. To address this challenge, variety of macromolecule delivery approaches or reagents have been developed. Our group has discovered and successfully employed small molecule DMSO (dimethyl sulfoxide) [11] and BIT (1,2-benzisothiazolin-3-one) [12] to facilitate the penetrating efficiency of TAT or TAT fused conjugates for a range of cell types, the unknown mechanism of enhancement and their potential side effects in high concentration may affect their application in clinic. We demonstrate that the system of CPP in hypertonic medium combined with osmoprotecants allows the highly efficient delivery of protein cargos
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