Delivery lysosomal

Several dendrimeric borane and carborane systems have been developed for use as boron-delivery agents.147 In vitro tests with a boronated dendrimer-epidermal growth factor bioconjugates indicated that they are endocytosed, resulting in the accumulation of boron in lysosomes. 

Other systems like electroporation have no lipids that might help in membrane sealing or fusion for direct transfer of the nucleic acid across membranes they have to generate transient pores, a process where efficiency is usually directly correlated with membrane destruction and cytotoxicity. Alternatively, like for the majority of polymer-based polyplexes, cellular uptake proceeds by clathrin- or caveolin-dependent and related endocytic pathways [152-156]. The polyplexes end up inside endosomes, and the membrane disruption happens in intracellular vesicles. It is noteworthy that several observed uptake processes may not be functional in delivery of bioactive material. Subsequent intracellular obstacles may render a specific pathway into a dead end [151, 154, 156]. With time, endosomal vesicles become slightly acidic (pH 5-6) and finally fuse with and mature into lysosomes. Therefore, polyplexes have to escape into the cytosol to avoid the nucleic acid-degrading lysosomal environment, and to deliver the therapeutic nucleic acid to the active site. Either the carrier polymer or a conjugated endosomolytic domain has to mediate this process [157], which involves local lipid membrane perturbation. Such a lipid membrane interaction could be a toxic event if occurring at the cell surface or mitochondrial membrane. Thus, polymers that show an endosome-specific membrane activity are favorable. 

Panyam J, Zhou WZ, Prabha S et al (2002) Rapid endo-lysosomal escape of poly(dl-lactide-co-glycolide) nanoparticles implications for drug and gene delivery. FASEB J 16 1217-1226... 

Human a-galactosidase is a 100 kDa homodimeric glycoprotein. Each 398 amino acid monomer displays a molecular mass of 45.3 kDa (excluding the glycocomponent) and is glycosylated at three positions (asparagines 108, 161 and 184). After administration (usually every second week by a 40 min infusion), the enzyme is taken up by various body cell types and directed to the lysosomes. This cellular uptake and delivery process appear to be mediated by mannose-... 

Figure 14.10 Overview of cellular entry of (non-viral) gene delivery systems, with subsequent plasmid relocation to the nucleus. The delivery systems (e.g. lipoplexes and polyplexes) initially enter the cell via endocytosis (the invagination of a small section of plasma membrane to form small membrane-bound vesicles termed endosomes). Endosomes subsequently fuse with golgi-derived vesicles, forming lysosomes. Golgi-derived hydrolytic lysosomal enzymes then degrade the lysosomal contents. A proportion of the plasmid DNA must escape lysosomal destruction via entry into the cytoplasm. Some plasmids subsequently enter the nucleus. Refer to text for further details...

The series of molecular events responsible for the uptake process constitutes the endocytic pathway, which enables cells to internalize macromolecules from the cell exterior, forming an endosome. The endosome is an intermediate organelle that serves as an essential component for many receptor-mediated signaling pathways and as a transport vector for eventual delivery to a specialized organelle known as the lysosome. Once in the lysosomal lumen, digestive enzymes provide essential metabolites from these macromolecules (i.e. free amino acids and lipids) directly to the cytosol for their use. 

Substrate availability to the cell is affected by the supply of raw materials from the environment. The plasma membranes of cells incorporate special and often specific transport proteins (translocases) or pores that permit the entry of substrates into the cell interior. Furthermore pathways in eukaryotic cells are often compartmentalized within cytoplasmic organelles by intracellular membranes. Thus we find particular pathways associated with the mitochondria, the lysosomes, the peroxisomes, the endoplasmic reticulum for example. Substrate utilization is limited therefore by its localization at the site of need within the cell and a particular substrate will be effectively concentrated within a particular organelle. The existence of membrane transport mechanisms is crucial in substrate delivery to, and availability at, the site of use. 

All in all, uptake studies with targeted systems carrying a pH-dependent fluorescent label in conjunction with monensin treatment at the end of the incubation period offer the possibility to discriminate between different intracellular trafficking pathways. At 37°C, an increasing fluorescence emission upon addition of monensin points toward accumulation of the targeted drug delivery system within acidic compartments of the cell, such as lysosomes. 

Wirth, M., Kneuer, C., Lehr, C.M., Gabor, F., Lectin-mediated drug delivery Discrimination between cytoadhesion and cytoinvasion and evidence for lysosomal accumulation of wheat germ agglutinin in the Caco-2 model. J Drug Target 10,... 

There are also multiple pathways for liposomes following cellular uptake. They may be delivered to lysosomes, recycled out of the cell, involved in transcytotic passage across an epithelial barrier, or delivered to other cellular compartments such as the Golgi network. Each route offers opportunities for selective delivery of macromolecular drugs and nanosized drugs so the need to comprehend endocytic pathways has never been more apparent (7). Figure 1 summarizes the different pathways of endocytosis. 

Both actin microfilaments and microtubules (see below) are involved in uptake as well as in the pathway to the lysosomes and the recycling pathway. Referring to clathrin-mediated uptake, Durrbach et al. showed actin participation in two steps of endocytosis the initial uptake mechanism and the delivery to lysosomes (116). 

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