Cell penetration enhancers

As mentioned earlier, penetration enhancers are extremely important for specific and rationally tailored delivery of pharmaceuticals. One novel class of such compounds are cell penetration enhancers (CPEs), which are unique peptides, first recognized in biological systems such as HIV, herpes enzymes, some fly [99] and frog species [100], and more [101]. These peptides have the ability to cause a perforation in the membranes of living cells thereby enhancing the penetration of molecules into the cells and target specific molecules to specific cells or nuclei [102, 103]. 

M. Cohen-Avrahami, A. Aserin, N. Garti, Hn mesophase and peptide cell-penetrating enhancers for improved transdermal delivery of sodium diclofenac. Colloids Surf. B 77, 131-138 (2010)... 

Sandri G, Rossi S, Ferrari F, Bonferoni MC, Zerrouk N, Caramella C (2004) Mucoadhesive and penetration enhancement properties of three different grades of hyaluronic acid using porcine buccal and vaginal tissue, Caco-2 cell lines and rat jejunum. J Pharm Pharmacol 56 1083-1090. 

Figure 2 Proposed pathways for liposomal entry into the cell enhanced by peptides. These include direct cell entry suggested as the mechanism of entry by cell-penetrating peptides and receptor-mediated endocytosis by caveolae- and clathrin-dependent endocytosis.

Marty C, Meylan C, Schott H, et al. Enhanced heparan sulfate proteoglycan-mediated uptake of cell-penetrating peptide-modified liposomes. Cell Mol Life Sci 2004 61 (14) 1785-1794. 

Irritation is veiy subjective and may differ widely from treatment to control subjects. Most irritation occurs as a result of penetration enhancers. Evaluation of toxicity and irritation should be concerned with mucosal tissue irritation, the extent of damage to the mucosal cells, and the rate of recovery. 

Yen, W.C., and Y.H.L. Lee. 1995. Penetration enhancement effect of Pz-peptide, a paracellularly transported peptide, in rabbit intestinal segments and Caco-2 cell monolayers. J Control Release 36 25. 

FIGURE 12.1 Penetration enhancer activity, (a) Action at intercellular lipids. Some of the ways by which penetration enhancers attack and modify the well-organized intercellular lipid domain of the stratum comeum. (b) Action at desmosomes and protein structures. Such dramatic disruption by enhancers (particularly potent solvents) as they split the stratum corneum into additional squames and individual cells would be clinically unacceptable, (c) Action within comeocytes. Swelling, further keratin denaturation and vacuolation within individual horny layer cells would not be so drastic but would usually be cosmetically challenging (see Menon and Lee [69] for further details). (Reproduced from Barry, B.W., Nat. Biotechnol. 22, 165, 2004. With permission.)... 

Penetration enhancers have been used to facilitate the absorption of higher molecular weight molecules. The mode of action of the surfactant enhancers is often attributed to membrane damage [37]. However, studies in epithelial cell monolayers suggest that some surfactant-based absorption enhancers act primarily by increasing the permeability of tight junctions [38]. Nevertheless, except for the chelators and nonsurfactants, which exert their... 

FIGURE 8.32 (a) A bright field epi-fluorescence image of two polymorphonuclear white cells (probably neutrophils), (b) The same cells shown in (a) imaged via the Hoechst stain. Note the differences in conformation between the two nuclei. The cell on the top is stuck at the entrance to a channel, and the cell at the bottom is being deformed into a channel. The channels are coated with polyurethane to reduce cell adhesion and to enhance white cell penetration [1175], Reprinted with permission from Springer Science and Business Media. 

However, as mentioned previously, a serious drawback associated with the use of penetration enhancers is their potential deleterious effect to the epithelial tissue, either directly, by damaging vital cell structures... 

Most compounds of interest for nasal delivery have a molecular weight in excess of 1,000 Da and until recently were thought to cross the cells endocytically. However, a recent study in rats has shown the transport of fluoroscein isothiocyanate (FITC)-labeled dextran (M. Wt. = 3,000 Da) to be via the paracellular pathway, with only a proportion moving endocytically. Hardly any transport of FITC-labeled dextran with a molecular weight of 10,000 Da was observed unless a penetration enhancer was coadministered, but the penetration enhancer, sodium taurodihydrofusidate (STDHF), caused cell swelling and extrusion of mucus. 

Already in 1965, Ryser and Hancock provided evidence that histones and polyamino acids could greatly enhance albumin uptake by cultured tumor cells (6). More recently, several polybasic peptides (so-called protein transduction domains, PTDs or cell-penetrating peptides, CPPs) have been shown to efficiently mediate uptake of nucleic acids, bioactive peptides, phage particles, and liposomes into a wide variety of mammalian cells. The initially proposed ability of CPPs to penetrate plasma membranes via a temperature-independent, non-endocytotic pathway was later shown to be a fixation artifact, and it is currently widely accepted that CPP-mediated macromolecular delivery follows energy-dependent endocytotic pathways that in most cases depend on the expression of cell-surface heparan sulfate proteoglycans (HSPGs) (7). 

Lee VHL, Yamamoto A, Kompella UB (1991) Mucosal penetration enhancers for facilitation of peptide and protein drug absorption. Crit Rev Ther Drug Carrier Syst 8 91-192 Matter K, Brauchbar M, Bucher K et al. (1990) Sorting of endo-geneous plasma membrane proteins occurs from two sites in cultured human intestinal epithelial cells, CACO-2. Cell 60 429-437... 

Table 1 Selected experimental dipeptide ICE/ced-3 family of cysteine proteases inhibitors having improved cell penetration and metabolic stability resulting in enhanced bioavailability. H-, 13C NMR, and mass spectral data supplied only for Entry 1 and its intermediates...

Various chitosan derivatives of enhanced solubility, mucoadhesive, and permeation properties were developed. V-Trimethyl chitosan chloride (TMC) is a quater-nized derivative of chitosan with superior aqueous solubility over a broader pH range and penetration-enhancing properties under physiological conditions [78]. Carboxymethylated chitosan (CMChi) is a polyampholytic polymer able to form viscoelastic gels in aqueous environments. CMChi appears to be less potent compared with the quaternized derivative. Neither TMC nor CMChi have been found to provoke damage of the cell membrane, and therefore, they should not alter the viability of nasal epithelial cells [79],... 

Penetration enhancers act by increasing the permeability of the corneal cell membrane and/or loosening the tight junctions between the epithelial cells, which primarily restrict the entry of molecules via the paracellular pathway. Classes of penetration enhancers include surfactants, bile salts, calcium chelators, preservatives, fatty acids, and some glycosides such a saponin. 

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