Main aim

The utilisation of macromolecules in therapy of cancer and other diseases is becoming increasingly relevant. Recent advances in molecular biology and biotechnology have made it possible to improve targeting and design of cytotoxic agents or DNA complexes for clinical applications. To achieve the expected biological effect of these macromolecules in many cases internalisation to the cell cytosol is crucial. At an intracellular level, the most fundamental obstruction for cytosolic release of the therapeutic molecule is the membrane-barrier of the endocytic vesicles. Photochemical internalisation (PCI) is a novel technology for release of endocytosed macromolecules into the cytosol. The technology is based on the use of photosensitizers located in endocytic vesicles that upon activation by light induce a release of macromolecules from their compartmentalization in endocytic vesicles. PCI has been shown to potentiate the biological activity of a large variety of macromolecules and other molecules that do not readily penetrate the plasma membrane, including proteins, peptides, peptide nucleic acids and DNA delivered as a complex with cationic polymers or incorporated in adenovirus. The group’s main focus is to develop PCI for therapeutic purposes.

Photodynamic therapy (PDT)

Porphyrins and many other photosensitizing compounds have been shown to induce cytotoxic effects on cells and tissues. These effects have been utilized to treat several types of neoplastic diseases. The treatment is named photodynamic therapy (PDT) and is based on injection of a photosensitizing and tumorlocalizing dye followed by exposure of the tumor region to light. The cytotoxic effect is mediated mainly through the formation of singlet oxygen. This reactive intermediate has a very short lifetime in cells (<0.04ms). Thus, the primary cytotoxic effect of PDT is executed during light exposure and very close to the sites of formation of singlet oxygen. Singlet oxygen reacts with and oxidizes proteins (histidine, tryptophan, methionine, cysteine, tyrosine), DNA (guanine), unsaturated fatty acids and cholesterol.

What is PCI?

The figure shows an illustration of how molecules can enter cytosol after pho-tochemical treatment.The photosensitizer (S) and the selected molecule (M) are endocytosed by the cells, ((I) illustrates the invagination of the plasma membrane) and both com-pounds ends up in the same vesicles (II). When these vesicles are exposed to light the membranes of these vesi-cles will be ruptured and the contents released (III).
The figure shows an illustration of how molecules can enter cytosol after pho-tochemical treatment.The photosensitizer (S) and the selected molecule (M) are endocytosed by the cells, ((I) illustrates the invagination of the plasma membrane) and both com-pounds ends up in the same vesicles (II). When these vesicles are exposed to light the membranes of these vesi-cles will be ruptured and the contents released (III).

Photochemical internalisation (PCI) is a unique novel technology (invented at the Norwegian Radium Hospital) that has emerged from photodynamic therapy (PDT). In PCI photochemical reactions are used to realise or potentiate the therapeutic effect of a lot of different molecules. These are molecules that are taken into the target cell by endocytosis and that have to be released from the endocytic vesicles to be able to exert a therapeutic effect. Examples of such molecules are: many proteins (e.g. several protein toxins), different types of oligonucleotides, and genes and viruses for gene therapy. Many of these molecules have a large therapeutic potential that it has so far not been possible to fully exploit. In the PCI technology light is used to relocate the therapeutic molecules from endocytic vesicles into the cell cytosol. In this way the molecules can reach their correct intracellular target, e.g. in the cell cytosol or nucleus, and exert a therapeutic effect that would otherwise been precluded by lysosomal degradation. It is well documented that a number of photosensitizers, including di- and tetrasulfonated aluminium phthalocyanine (AlPcSn) and sulfonated tetraphenylporphines (TPPSn), are located in endosomes and lysosomes. The introduction of molecules into the cytosol is achieved by first exposing the cells or tissue to a photosensitizing dye and the molecule which one wants to deliver, both of which should preferentially localize in endosomes and/or lysosomes. Secondly, the cells or tissues are exposed to light of wavelengths inducing a photochemical reaction. This photochemical reaction will lead to disruption of lysosomal and/or endosomal membranes and the contents of these vesicles will be released into the cytosol. The principle is illustrated in the figure.

In vitro PCI, has been documented efficient in internalization of type I ribosome-inactivating plant toxins (RIPs), an immunotoxin, a peptide (for use in cancer vaccine), horseradish peroxidase, ribozymes, oligodeoxynucleotides, plasmids and an adenovirus gene therapy vector.

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