" Mechanisms involved in d-aminolevulinic acid (ALA)-induced photosensitivity of tumor cells: Relation of ferrochelatase and uptake of ALA to the accumulation of protoporphyrin Yoshiko Ohgari a , Yuki Nakayasu a , Sakihito Kitajima a , Mari Sawamoto a , Hajime Mori a,b , Osamu Shimokawa c , Hirofumi Matsui c , Shigeru Taketani a,b, * a Department of Biotechnology, Kyoto Institute of Technology, Kyoto 606-8585, Japan b Insect Biomedical Center, Kyoto Institute of Technology, Kyoto 606-8585, Japan c Graduate School of Comprehensive Human Sciences, Tsukuba University, Tukuba 305-8575, Japan biochemical pharmacology 71 (2005) 42–49 article info Article history: Received 31 August 2005 Accepted 7 October 2005 Keywords: Ferrochelatase d-Aminolevlinic acid Protoporphyrin Photo-damage PDT Abbreviations: ALA, d-aminolevlinic acid DFO, desferrioxamine DMEM, Dulbecco’s modified Eagle’s medium FCS, fetal calf serum HO-1, heme oxygenase-1 PBG, phorphobilinogen PDT, Photodynamic therapy PVDF, poly(vinylidene difluoride) SDS-PAGE, sodium dodecylsulfate-polyacrylamide gel electrophoresis abstract Photodynamic therapy (PDT) using d-aminolevulinic acid (ALA)-induced accumulation of protoporphyrin IX is a useful approach to the early detection and treatment of cancers. To investigate the role of ferrochelatase in the accumulation of protoporphyrin, we first made mouse fibroblast Balb/3T3 cells highly expressing ferrochelatase and examined the ALA- induced photo-damage as well as the accumulation of porphyrin in the cells. When the ferrochelatase-transfected cells were treated with ALA and then exposed to visible light, they became resistant to the light without accumulating porphyrins, with a concomitant increase in the formation of heme. The accumulation of protoporphyrin was also abolished in human erythroleukemia K562 cells stably expressing mouse ferrochelatase. When mouse fibrosarcoma MethA cells, mouse fibroblast L929 cells and Balb/3T3 cells were treated with ALA, the greatest accumulation of protoporphyrin and the greatest level of cell death in response to the light were observed in MethA cells. The expression level of ferrochelatase was the lowest in MethA cells, while that of porphobilinogen deaminase was similar among all three cell lines. Moreover, an iron-chelator, desferrioxamine, which sequesters iron preventing the ferrochelatase reaction, enhanced the photo-damage as well as the accu- mulation of protoporphyrin in ALA-treated L929 cells. Thus, the light-induced cell death was tightly coupled with the accumulation of protoporphyrin caused by a decrease in ferroche- latase. Finally, we examined the uptake of ALA by MethA, L929 and Balb/3T3 cells. The extent of the uptake by MethA and L929 cells was greater, indicating a greater accumulation of protoporphyrin than in the Balb/3T3 cells. Taken together, not only the low level of ferrochelatase but also the augmented uptake of ALA contributes to the ALA-induced accumulation of protoporphyrin IX and subsequent photo-damage in cancer cells. # 2005 Elsevier Inc. All rights reserved. * Corresponding author. Tel.: +81 75 724 7789; fax: +81 75 724 7760. E-mail address: taketani@kit.ac.jp (S. Taketani). available at www.sciencedirect.com journal homepage: www.elsevier.com/locate/biochempharm 0006-2952/$ – see front matter # 2005 Elsevier Inc. All rights reserved. doi:10.1016/j.bcp.2005.10.019