Protoporphyrin IX Formation Research Articles

Effects of the inhibitors of energy metabolism, lonidamine and levamisole, on 5-aminolevulinic-acid-induced photochemotherapy.

The ability of endogenously synthesized protoporphyrin IX (PpIX) to damage Chinese hamster lung fibroblasts of the line V79 by exposure to light was examined. This treatment induced reduction of cellular ATP, GTP, of the NADH/NAD+ ratio and of oxygen consumption. The present results indicate a close relationship between inhibition of respiration of irradiated cells and their ability to survive, e.g. 1 min of light exposure induced 90% inhibition of oxygen consumption and inactivation of approximately 95% of the cells, while the cellular content of ATP was reduced by only 15%. This indicates that the mitochondria are one of the primary targets of 5-aminolevulinic acid (ALA)-mediated photochemotherapy (PCT). In the present study, ALA-PCT was combined with the modulators of the glycolysis and the respiration chain, levamisole (LEV) and lonidamine (LND). A synergistic effect of combining ALA-PCT with non-toxic concentrations of LND was observed when LND was given prior to light exposure. This synergism was observed despite a substantial LND-induced inhibition of PpIX formation. At increasing doses of LND (>0.15 mM) the combination treatment becomes less efficient. This is due to the inhibition of PpIX synthesis induced by LND. A synergistic effect of ALA-PDT and LEV was found when LEV was given prior to light exposure. This was at least partly due to an LEV-stimulated effect on ALA-induced PpIX formation. However, it is not clear from the present results whether LEV may perturb energy metabolism in V79 cells since LEV alone did not reduce the energy charge or the NADH/NAD+ ratio. When LEV or LND were given after ALA-PCT, these 2 treatment modalities acted in an additive or slightly synergistic manner.

Does δ-aminolaevulinic acid induce genotoxic effects?

5-Aminolaevulinic acid (ALA) is a precursor of protoporphyrin IX (PpIX) in the biosynthetic pathway for haem. The presence of exogenous ALA bypasses the feedback control and may induce the accumulation of PpIX. Since haem-containing enzymes are essential for energy metabolism, every nucleated cell in the body must have at least a minimal capacity to synthesize PpIX. Photodynamic therapy (PDT), which is the treatment of malignant lesions with light following the administration of a tumour-localizing photosensitizers leads to oxidative damage, including the formation of genotoxic membrane degradation products via lipid peroxidation. In addition, it has been demonstrated that ALA itself can form the reactive oxygen species O 2, H 2O 2 and OH − by auto-oxidation, suggesting that it could potentially induce DNA damage. Therefore cultures of rat hepatocytes, which have been demonstrated to be very sensitive indicators for genotoxic effects induced by the lipid peroxidation product 4-hydroxynonenal and analogous aldehydes, were examined for possible mutagenic effects after treatment with ALA in the absence of light. The cytogenetic endpoints determined were chromosomal aberrations and the induction of micronuclei. Compared with the controls, significantly elevated levels of chromosomal aberrations and micronuclei were observed at concentrations of 1 μg ml 1 or greater. Chromosomal aberrations and micronuclei were found to increase up to a concentration of 100 μg ml 1 ALA. While micronuclei decrease at higher concentrations, chromosomal aberrations remain at the same level. The kinetics of PpIX formation after induction with 100 and 1000 μg ml −1 ALA appear to be the same for both concentrations, suggesting that the induction of chromosomal aberrations may be due to PpIX.