Delivery of nucleic acids and nanomaterials by cell-penetrating peptides: opportunities and challenges

Abstract

Many viral and nonviral systems have been developed to aid delivery of biologically active molecules into cells. Among these, cell-penetrating peptides (CPPs) have received increasing attention in the past two decades for biomedical applications. In this review, we focus on opportunities and challenges associated with CPP delivery of nucleic acids and nanomaterials. We first describe the nature of versatile CPPs and their interactions with various types of cargoes. We then discuss in vivo and in vitro delivery of nucleic acids and nanomaterials by CPPs. Studies on the mechanisms of cellular entry and limitations in the methods used are detailed.

Figure 1

Cell-penetrating peptides as a tool to deliver biologically active molecules.

Figure 2

Reaction scheme for linking CPPs to cargoes. The cargoes can be linked to the CPPs through a covalent linkage method such as (a) bissulfosuccinimidyl suberate, (b) carbodiimide, or (c) Sulfo-SMCC with a cysteine-modified CPP, or through a noncovalent method such as (d) biotin-streptavidin interaction.

Figure 3

Simplified conceptual diagram (not drawn proportionally in size) of exogenous siRNA-mediated gene silencing. (a) The siRNA (usually small hairpin RNA, shRNA) can be modified to covalently interact with CPPs and then be transported through the cell membrane. (b) shRNA binds to the double-strand RNA binding domain (dsRBD) of the enzyme Dicer and then is processed. (c) The processed RNA is incorporated into the RNA-induced silencing complex (RISC). The passenger strand RNA is degraded. (d) The guide strand RNA along with the RISC binds to a complementary sequence of a targeted mRNA. (e) The targeted mRNA is degraded and translation disrupted.

Figure 4

(a) Synthesis of water-soluble carboxylated CdSe/ZnS quantum dots. Upon addition of ZnS as a shell to protect Cd core, the surface was modified with 1, 2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[carboxy(polyethyleneglycol)-2000] (DSPE-PEG 2000) and 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethyleneglycol)-2000] (PEG-2 PE). The amount and ratio of PEG2-PE and DSPE-PEG(200) determine suspension stability in water. (b) Fluorescence of CdSe/ZnS quantum dot in live cells with (left) and without (right) nona-arginine after a 1-hour exposure [17].

Figure 5

HR9 CPP facilitates cellular uptake of green fluorescent nanodiamonds.

Figure 6

Diagram illustrating a comprehensive workflow of experiments designed to characterize the cellular uptake, intracellular uptake, and subcellular localization of CPPs and their cargoes.

Figure 7

Comparisons of clathrin- and caveolin-dependent cellular uptake pathways using pharmacological inhibitors and RNAi technique [17].