Cell penetration: scope and limitations by the application of cell-penetrating peptides.

The penetration of polar or badly soluble compounds through a cell membrane into live cells requires mechanical support or chemical helpers. Cell-penetrating peptides (CPPs) are very promising chemical helpers. Because of their low cytotoxicity and final degradation to amino acids, they are particularly favored in in vivo studies and for clinical applications. Clearly, the future of CPP research is bright; however, the required optimization studies for each drug require considerable individualized attention. Thus, CPPs are not the philosopher's stone. As of today, a large number of such transporter peptides with very different sequences have been identified. These have different uptake mechanisms and can transport different cargos. Intracellular concentrations of cargos can reach a low micromole range and are able to influence intracellular reactions. Internalized ribonucleic acids such as small interfering RNA (siRNA) and mimics of RNA such as peptide nucleic acids, morpholino nucleic acids, and triesters of oligonucleotides can influence transcription and translation. Despite the highly efficient internalization of antibodies, enzymes, and other protein factors, as well as siRNA and RNA mimics, the uptake and stabile insertion of DNA into the genome of the host cells remain substantially challenging. This review describes a wide array of differing CPPs, cargos, cell lines, and tissues. The application of CPPs is compared with electroporation, magnetofection, lipofection, viral vectors, dendrimers, and nanoparticles, including commercially available products. The limitations of CPPs include low cell and tissue selectivity of the first generation and the necessity for formation of fusion proteins, conjugates, or noncovalent complexes to different cargos and of cargo release from intracellular vesicles. Furthermore, the noncovalent complexes require a strong molar excess of CPPs, and extensive experimentation is required to determine the most optimal CPP for any given cargo and cell type. Yet to predict which CPP is optimal for any given target remains a complex question. More recently, there have been promising developments: the enhancement of cell specificity using activatable CPPs, specific transport into cell organelles by insertion of corresponding localization sequences, and the transport of drugs through blood-brain barriers, through the conjunctiva of eyes, skin, and into nerve cells. Proteins, siRNA, and mimics of oligonucleotides can be efficiently transported into cells and have been tested for treatment of certain diseases. The recent state of the art in CPP research is discussed together with the overall scope, limitations, and some recommendations for future research directions.

Reissmann S 《-》

Peptide vectors for the nonviral delivery of nucleic acids.

Hoyer J ，Neundorf I 《-》

A survey on "Trojan Horse" peptides: opportunities, issues and controlled entry to "Troy".

Cell-penetrating peptides (CPPs), often vividly termed as the "Trojan Horse" peptides, have attracted considerable interest for the intracellular delivery of a wide range of cargoes, such as small molecules, peptides, proteins, nucleic acids, contrast agents, nanocarriers and so on. Some preclinical and clinical developments of CPP conjugates demonstrate their promise as therapeutic agents for drug discovery. There is increasing evidence to suggest that CPPs have the potential to cross several bio-barriers (e.g., blood-brain barriers, intestinal mucosa, nasal mucosa and skin barriers). Despite revolutionary process in many aspects, there are a lot of basic issues unclear for these entities, such as internalization mechanisms, translocation efficiency, translocation kinetics, metabolic degradation, toxicity, side effect, distribution and non-specificity. Among them, non-specificity remains a major drawback for the in vivo application of CPPs in the targeted delivery of cargoes. So far, diverse organelle-specific CPPs or controlled delivery strategies have emerged and improved their specificity. In this review, we will look at the opportunities of CPPs in clinical development, bio-barriers penetration and nanocarriers delivery. Then, a series of basic problems of CPPs will be discussed. Finally, this paper will highlight the use of various controlled strategies in the organelle-specific delivery and targeted delivery of CPPs. The purpose of this review will be to emphasize most influential advance in this field and present a fundamental understanding for challenges and utilizations of CPPs. This will accelerate their translation as efficient vectors from the in vitro setting into the clinic arena, and retrieve the entry art to "Troy".

Shi NQ ，Qi XR ，Xiang B ，Zhang Y ... -  《-》

Cell penetrating peptides: the potent multi-cargo intracellular carriers.

Kardani K ，Milani A ，H Shabani S ，Bolhassani A ... -  《-》

Transduction of peptides and proteins into live cells by cell penetrating peptides.

Internalization of peptides and proteins into live cells is an essential prerequisite for studies on intracellular signal pathways, for treatment of certain microbial diseases and for signal transduction therapy, especially for cancer treatment. Cell penetrating peptides (CPPs) facilitate the transport of cargo-proteins through the cell membrane into live cells. CPPs which allow formation of non-covalent complexes with the cargo are used primarily in this study due to the relatively easy handling procedure. Efficiency of the protein uptake is estimated qualitatively by fluorescence microscopy and quantitatively by SDS-PAGE. Using the CPP cocktail JBS-Proteoducin, the intracellular concentrations of a secondary antibody and bovine serum albumin can reach the micromolar range. Internalization of antibodies allows mediation of intracellular pathways including knock down of signal transduction. The high specificity and affinity of antibodies makes them potentially more powerful than siRNA. Thus, CPPs represent a significant new possibility to study signal transduction processes in competition or in comparison to the commonly used other techniques. To estimate the highest attainable intracellular concentrations of cargo proteins, the CPPs are tested for cytotoxicity. Cell viability and membrane integrity relative to concentration of CPPs are investigated. Viability as estimated by the reductive activity of mitochondria (MTT-test) is more sensitive to higher concentrations of CPPs versus membrane integrity, as measured by the release of dead cell protease. Distinct differences in uptake efficiency and cytotoxic effects are found using six different CPPs and six different adhesion and suspension cell lines.

Mussbach F ，Franke M ，Zoch A ，Schaefer B ，Reissmann S ... -  《-》