Cell-penetrating peptides: mechanism and kinetics of cargo delivery
Introduction
Over the last 10 years, it has been found that certain peptides and proteins can penetrate the cell membrane and enter the cell. A variety of cargo molecules can be attached to these peptides and proteins and translocated into the cell. Carrier peptides and proteins thus constitute a new class of potential drug delivery vectors. Currently, about hundred such peptides and a few proteins are known. In this review, we shall concentrate on the carrier peptides and aim to give an up to date survey of their structure, kinetics of cell penetration, cargo delivery potential, and undesirable side effects. Transduction proteins will be only briefly mentioned here, since they are covered in a separate review. However, in most cases only fractions of the transduction protein sequence are necessary for translocation and internalization of cargo into the cells, so these peptides will be included in the review.
Membranes of eucaryotic cells and organelles, as well as the cell wall and membrane of pathogenic microorganisms, constitute a serious barrier for the access of hydrophilic drugs to their target molecules inside the cell structures. To overcome problems of conventional and gene drug delivery, various techniques have been developed. A conventional procedure for delivering genetic material is to use viral vectors, but treating genetic disorders with this method has met with only limited success [1]. Alternative non-viral methods, such as electroporation, microinjection, and the use of liposomes, have been developed for conventional and gene drug delivery. These methods have been proved to be effective in vitro and for research purposes, but show limited potential for delivery in vivo due to toxicity, cell damage, and immunogenicity. They are also technically demanding in their application, lack tissue and cell specificity, and can deliver material to only a limited amount of cells. In view of these considerations, transduction peptides and proteins offer a promising new tool for noninvasive delivery of hydrophilic drugs and genetic material.
Section snippets
Terminology, classification, and structure
A unified terminology and classification of the carrier peptides has not yet been developed. The commonly used name is “cell-penetrating peptide” (CPP) which was more widely introduced in a review article [2] and in the first book [3] on this subject. Very generally, CPPs are up to 30 amino acid amphiphilic peptides, which can be internalized by cells by mechanisms that require no energy and are receptor mediated or not. According to a recently suggested classification, CPPs can be arranged in
Monitoring internalization of CPPs and cargo
CPPs cannot usually be observed directly and must be appropriately labelled to allow reliable detection of the small amounts that are usually internalized. Labelled CPP or cargo can be detected by radioactivity, fluorescence emission, fluorescence quenching, specific labelling with dyes or by enzymatic activity of the cargo. Some of the detection methods are very convenient for visualizing CPP internalized in cells but do not have a sufficiently rapid response time for kinetic studies. An
Conclusions and perspectives
The large number of different CPPs synthesized so far have been demonstrated to enter different cells, tissues and organs. They are able to promote translocation of various types of useful cargo, ranging from small molecules to proteins and large supramolecular particles, with great efficiency and reasonable velocity. Delivery of cargo has been achieved also into some systems that are usually difficult to cross, for instance the blood–brain barrier. As described above, CPPs have already been
Acknowledgment
This work was supported by grants from European Community (QLK3-CT-2002-01989), Swedish Research Councils (VR-NT and VR-Med), and Ministry of Education, Research and Sport of the Republic of Slovenia (PO-0505-0381).
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