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Advanced Drug Delivery Reviews (v.57, #4)
Protein- and peptide-mediated transduction: mechanisms and implications for drug delivery
by Vladimir P. Torchilin Theme Editor (pp. 489-490).
Protein transduction: from physiology to technology and vice versa
by Alain Prochiantz (pp. 491-493).
Adaptive translocation: the role of hydrogen bonding and membrane potential in the uptake of guanidinium-rich transporters into cells by Jonathan B. Rothbard; Theodore C. Jessop; Paul A. Wender (pp. 495-504).
A mechanistic hypothesis is presented for how water-soluble guanidinium-rich transporters attached to small cargoes (MW ca. <3000) can migrate across the non-polar lipid membrane of a cell and enter the cytosol. Positively charged and water-soluble, arginine oligomers can associate with negatively charged, bidentate hydrogen bond acceptor groups of endogenous membrane constituents, leading to the formation of membrane-soluble ion pair complexes. The resultant less polar, ion pair complexes partition into the lipid bilayer and migrate in a direction, and with a rate, influenced by the membrane potential. The complex dissociates on the inner leaf of the membrane and the transporter conjugate enters the cytosol. This mechanism could also be involved in the translocation of guanidinium-rich molecules that are endocytosed due to their size or the conditions of the assay, across the endosomal membrane.
Keywords: Arginine-rich transporters; Membrane potential; Ion pairs; Drug delivery; Protein translocation; Endocytosis; HIV tat; Antennapedia; FGF
Nuclear delivery of macromolecules: barriers and carriers by Mattias Belting; Staffan Sandgren; Anders Wittrup (pp. 505-527).
Recent evidence for efficient delivery of macromolecules, such as peptides and nucleic acids, from the cell exterior to the nucleus offers the interesting possibility of developing novel treatments directed at intranuclear targets. The findings should also stimulate the search for physiological ligands that utilize similar transport mechanisms to regulate pathobiological processes. Cytokines, growth factors and their receptors, as well as morphogens have all been shown to enter the nucleus to evoke biological responses in target cells. The rational design of intracellular drug delivery vehicles requires an increased understanding of the elaborate systems that mediate cellular communication and coordination with the extracellular environment without inflicting on the integrity of the cell. This review discusses some aspects of the carriers and barriers in macromolecular transport.
Keywords: Gene transfer; Endocytosis; Virus; Proteoglycan; Nucleic acid; Polybasic peptide; Drug delivery
Cell-penetrating peptides: mechanism and kinetics of cargo delivery by Matjaž Zorko; Ülo Langel (pp. 529-545).
Cell-penetrating peptides (CPPs) are short peptides of less than 30 amino acids that are able to penetrate cell membranes and translocate different cargoes into cells. The only common feature of these peptides appears to be that they are amphipathic and net positively charged. The mechanism of cell translocation is not known but it is apparently receptor and energy independent although, in certain cases, translocation can be partially mediated by endocytosis. Cargoes that are successfully internalized by CPPs range from small molecules to proteins and supramolecular particles. Most CPPs are inert or have very limited side effects. Their penetration into cells is rapid and initially first-order, with half-times from 5 to 20 min. The size of smaller cargoes does not affect the rate of internalization, but with larger cargoes, the rate is substantially decreased. CPPs are novel vehicles for the translocation of cargo into cells, whose properties make them potential drug delivery agents, of interest for future use.
Keywords: Drug delivery; Membrane translocation; Cellular uptake; Endocytosis; Toxicity
Membrane-permeable arginine-rich peptides and the translocation mechanisms by Shiroh Futaki (pp. 547-558).
The intracellular delivery of proteins and other bioactive molecules using membrane-permeable carrier peptide vectors opens the possibility of establishing novel methods of elucidating and controlling cell functions with therapeutic potentials. One of the most typical peptide vectors is a short, arginine-rich peptide segment derived from the human immunodeficiency virus (HIV)-1 Tat protein. We have shown that not only the Tat peptide, but also various arginine-rich oligopeptides possess very similar characteristics in translocation and abilities as a delivery vector. This review summarizes the structures of these peptide vectors, especially the Tat and other arginine-rich peptides, and the current understanding of their internalization mechanisms.
Keywords: HIV-1 Tat; Oligoarginine; Penetratin; Cell permeable peptide; Protein transduction; Drug delivery
Tat peptide-mediated cellular delivery: back to basics by Hilary Brooks; Bernard Lebleu; Eric Vivès (pp. 559-577).
Peptides are emerging as attractive drug delivery tools. The HIV Tat-derived peptide is a small basic peptide that has been successfully shown to deliver a large variety of cargoes, from small particles to proteins, peptides and nucleic acids. The ‘transduction domain’ or region conveying the cell penetrating properties appears to be confined to a small (9 amino acids) stretch of basic amino acids, with the sequence RKKRRQRRR [S. Ruben, A. Perkins, R. Purcell, K. Joung, R. Sia, R. Burghoff, W.A. Haseltine, C.A. Rosen, Structural and functional characterization of human immunodeficiency virus tat protein, J. Virol. 63 (1989) 1–8; S. Fawell, J. Seery, Y. Daikh, C. Moore, L.L. Chen, B. Pepinsky, J. Barsoum, Tat-mediated delivery of heterologous proteins into cells, Proc. Natl. Acad. Sci. U. S. A. 91 (1994) 664–668; E. Vives, P. Brodin, B. Lebleu, A truncated HIV-1 Tat protein basic domain rapidly translocates through the plasma membrane and accumulates in the cell nucleus, J. Biol. Chem. 272 (1997) 16010–16017; S. Futaki, T. Suzuki, W. Ohashi, T. Yagami, S. Tanaka, K. Ueda, Y. Sugiura, Arginine-rich peptides. An abundant source of membrane-permeable peptides having potential as carriers for intracellular protein delivery, J. Biol. Chem. 276 (2001) 5836–5840.]. The mechanism by which the Tat peptide adheres to, and crosses, the plasma membrane of cells is currently a topic of heated discussion in the literature, with varied findings being reported. This review aims to bring together some of those findings. Peptide interactions at the cell surface, and possible mechanisms of entry, will be discussed together with the effects of modifying the basic sequence and attaching a cargo.
Keywords: TAT; Uptake; Cell delivery; CPP
Transmembrane delivery of protein and peptide drugs by TAT-mediated transduction in the treatment of cancer by Jehangir S. Wadia; Steven F. Dowdy (pp. 579-596).
The direct intracellular delivery of proteins, or active peptide domains, has, until recently, been difficult to achieve due primarily to the bioavailability barrier of the plasma membrane, which effectively prevents the uptake of macromolecules by limiting their passive entry. Traditional approaches to modulate protein function have largely relied on the serendipitous discovery of specific drugs and small molecules which could be delivered easily into the cell. However, the usefulness of these pharmacological agents is limited by their tissue distribution and unlike ‘information-rich’ macromolecules, they often suffer from poor target specificity, unwanted side-effects, and toxicity. Likewise, the development of molecular techniques, over the past several decades, for gene delivery and expression of proteins has provided for tremendous advances in our understanding of cellular processes but has been of surprisingly little benefit for the management of genetic disorders. Apart from these gains however, the transfer of genetic material into eukaryotic cells either using viral vectors or by non-viral mechanisms such as microinjection, electroporation, or chemical transfection remains problematic. Moreover, in vivo, gene therapy approaches relying on adenoviral vectors are associated with significant difficulties relating to a lack of target specificity and toxicity which have contributed to poor performance in several clinical trials. Remarkably, the recent identification of a particular group of proteins with enhanced ability to cross the plasma membrane in a receptor-independent fashion has led to the discovery of a class of protein domains with cell membrane penetrating properties. The fusion of these protein transduction domain peptide sequences with heterologous proteins is sufficient to cause their rapid transduction into a variety of different cells in a rapid, concentration-dependent manner. Moreover, this novel technique for protein and peptide delivery appears to circumvent many problems associated with DNA and drug based methods. This technique may represent the next paradigm in our ability to modulate cell function and offers a unique avenue for the treatment of disease.
Keywords: HIV1 TAT; Protein transduction; Cancer; Protein therapy; Peptide therapy
Transcellular protein transduction using the Tat protein of HIV-1 by Antonio Fittipaldi; Mauro Giacca (pp. 597-608).
The Tat protein of HIV-1 is a powerful transactivator of gene expression. By interacting with a structured RNA sequence at the 5′ end of the viral mRNA, it promotes the remodeling of chromatin and the recruitment of processive RNA polymerase complexes at the viral promoter. In addition to these transcriptional functions, a short amino acid motif, highly enriched in basic amino acids, promotes the export of the protein from the expressing cells. Once in the extracellular environment, the same basic domain of Tat binds to cell surface heparan sulfate proteoglycans; through this interaction, the protein is internalized by a variety of different cell types. Cellular internalization of Tat and Tat fusion proteins requires the integrity of cell membrane lipid rafts and mainly occurs through caveolar endocytosis. The Tat basic domain, when attached to large protein cargos, also mediates their efficient cellular internalization and can be thus utilized for transcellular protein transduction. This property has already been successfully exploited for the delivery of heterologous proteins, nanoparticles, liposomes, phage and viral vectors, and plasmid DNA. The biological significance of intercellular Tat trafficking in the context of viral infection still remains elusive.
Keywords: Encocytosis; Human immunodeficiency virus type 1 (HIV-1); Lipid rafts; Protein transduction; Transactivation
Intracellular targeting of polymer-bound drugs for cancer chemotherapy by Aparna Nori; JindÅ™ich KopeÄ?ek (pp. 609-636).
Macromolecules have been traditionally employed as drug carriers due to their ability to selectively accumulate in malignant tissues compared to healthy tissues by either passive or active targeting, thus precluding undesirable side effects generated by free drug. The therapeutic activity proffered by such conjugates requires that the drug concentrate at its specific subcellular target such as the nucleus. Thus, the suitability of macromolecules as carriers also extends to their propensity to deliver the drug to a predetermined intracellular location. As binding a macromolecule to a drug facilitates cellular uptake by endocytosis, various approaches have been employed to either guide the drug to targets different from endosomal/lysosomal compartments by mediating vesicular escape, or to directly accomplish intracellular (cytoplasmic and nuclear) localization. This review discusses the utility of macromolecules in drug delivery and describes the numerous modalities (with a focus on cell-penetrating peptides) currently available for achieving effective intracellular drug delivery.
Keywords: Polymers; HPMA; Molecular signaling; Cell-penetrating peptides; Subcellular trafficking; Nuclear localization sequences
Intracellular delivery of large molecules and small particles by cell-penetrating proteins and peptides by Bhawna Gupta; Tatiana S. Levchenko; Vladimir P. Torchilin (pp. 637-651).
Cell-penetrating peptides (CPPs) have been used to overcome the lipophilic barrier of the cellular membranes and deliver large molecules and even small particles inside the cell for their biological actions. CPPs are being used to deliver inside cell a large variety of cargoes such as proteins, DNA, antibodies, contrast (imaging) agents, toxins, and nanoparticular drug carriers including liposomes. In this paper, we have reviewed the delivery of different molecules and particles mediated by TAT, Antp, VP22, and other CPPs as well as potential applications of these delivery systems in different areas of vaccine development, cancer immunotherapy, gene delivery, and cellular imaging.
Keywords: Intracellular delivery; Cell-penetrating peptides; Fusion proteins; Gene delivery; Toxins; Imaging agents; Particles; Liposomes
The role of alpha-helical structure in p53 peptides as a determinant for their mechanism of cell death: necrosis versus apoptosis by Ramon Rosal; Paul Brandt-Rauf; Matthew R. Pincus; Hsin Wang; Yuehua Mao; Yin Li; Robert L. Fine (pp. 653-660).
Peptides derived from the N-terminal and C-terminal regions of the p53 tumor suppressor protein, linked to the membrane transduction domain of Antennapedia, have both been found to have significant cytotoxic effects selectively in human cancer cells. However, the N-terminal and C-terminal p53 peptides apparently display very different mechanisms for their anticancer effects. These differential effects can be attributed to dissimilar abilities to form distinctive 3-dimensional structures in extracellular-matrix-like aqueous solution that enable unique and selective cancer cell membrane penetration and effect. N-terminally based p53 peptides, with their ability to form distinctive S-shaped helix–loop–helix structures, are able to rapidly disrupt cancer cell membranes via toroidal-like pore formation causing necrosis; conversely, C-terminally based p53 peptides, due to their more random coil configuration, can be transduced across cancer cell membranes and bind to its intracellular target to cause a Fas pathway mechanism of apoptosis.
Keywords: p53; Necrosis; Apoptosis; Peptides; NMR solution structure; Alpha helical