Skip to main content

Advertisement

SpringerLink
Log in

Serum-dependent export of protoporphyrin IX by ATP-binding cassette transporter G2 in T24 cells

  • Published:
Molecular and Cellular Biochemistry Aims and scope Submit manuscript

Abstract

Accumulation of protoporphyrin IX (PpIX) in cancer cells is a basis of 5-aminolevulinic acid (ALA)-induced photodymanic therapy. We studied factors that affect PpIX accumulation in human urothelial carcinoma cell line T24, with particular emphasis on ATP-binding cassette transporter G2 (ABCG2) and serum in the medium. When the medium had no fetal bovine serum (FBS), ALA induced PpIX accumulation in a time- and ALA concentration-dependent manner. Inhibition of heme-synthesizing enzyme, ferrochelatase, by nitric oxide donor (Noc18) or deferoxamine resulted in a substantial increase in the cellular PpIX accumulation, whereas ABCG2 inhibition by fumitremorgin C or verapamil induced a slight PpIX increase. When the medium was added with FBS, cellular accumulation of PpIX stopped at a lower level with an increase of PpIX in the medium, which suggested PpIX efflux. ABCG2 inhibitors restored the cellular PpIX level to that of FBS(–) samples, whereas ferrochelatase inhibitors had little effects. Bovine serum albumin showed similar effects to FBS. Fluorescence microscopic observation revealed that inhibitors of ABC transporter affected the intracellular distribution of PpIX. These results indicated that ABCG2-mediated PpIX efflux was a major factor that prevented PpIX accumulation in cancer cells in the presence of serum. Inhibition of ABCG2 transporter system could be a new target for the improvement of photodynamic therapy.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8
Fig. 9
Fig. 10

Similar content being viewed by others

Abbreviations

ABCG2:

ATP-binding cassette transporter G2

ALA:

5-Aminolevulinic acid

DFX:

Deferoxamine

FBS:

Fetal bovine serum

FTC:

Fumitremorgin C

NAO:

10-Nonyl acridine orange

Noc18:

1-Hydroxy-2-oxo-3,3-bis(2-aminoethyl)-1-triazene

PBS:

Phosphate-buffered saline

PDT:

Photodynamic therapy

PI:

Propidium iodide

PpIX:

Protoporphyrin IX

SA:

Succinyl acetone

References

  1. Peng Q, Warloe T, Berg K, Moan J, Kongshaug M, Giercksky KE, Nesland JM (1997) 5-Aminolevulinic acid-based photodynamic therapy. Clinical research and future challenges. Cancer 79:2282–2308

    Article  PubMed  CAS  Google Scholar 

  2. Datta SN, Loh CS, MacRobert AJ, Whatley SD, Matthews PN (1998) Quantitative studies of the kinetics of 5-aminolaevulinic acid-induced fluorescence in bladder transitional cell carcinoma. Br J Cancer 78:1113–1118

    Article  PubMed  CAS  Google Scholar 

  3. Hinnen P, de Rooij FW, van Velthuysen ML, Edixhoven A, van Hillegersberg R, Tilanus HW, Wilson JH, Siersema PD (1998) Biochemical basis of 5-aminolaevulinic acid-induced protoporphyrin IX accumulation: a study in patients with (pre)malignant lesions of the oesophagus. Br J Cancer 78:679–682

    Article  PubMed  CAS  Google Scholar 

  4. Krishnamurthy P, Xie T, Schuetz JD (2007) The role of transporters in cellular heme and porphyrin homeostasis. Pharmacol Ther 114:345–358. doi:10.1016/j.pharmthera.2007.02.001

    Article  PubMed  CAS  Google Scholar 

  5. Zutz A, Gompf S, Schägger H, Tampé R (2009) Mitochondrial ABC proteins in health and disease. Biochim Biophys Acta 1787:681–690. doi:10.1016/j.bbabio.2009.02.009

    Article  PubMed  CAS  Google Scholar 

  6. Ishikawa T, Nakagawa H, Hagiya Y, Nonoguchi N, Miyatake S, Kuroiwa T (2010) Key role of human ABC transporter ABCG2 in photodynamic therapy and photodynamic diagnosis. Adv Pharmacol Sci 2010:587306. doi:10.1155/2010/587306

    PubMed  Google Scholar 

  7. Schoenfeld N, Mamet R, Leibovici L, Epstein O, Teitz Y, Atsmon A (1988) Growth rate determines activity of porphobilinogen deaminase both in nonmalignant and malignant cell lines. Biochem Med Metab Biol 40:213–217

    Article  PubMed  CAS  Google Scholar 

  8. Krieg RC, Fickweiler S, Wolfbeis OS, Knuechel R (2000) Cell-type specific protoporphyrin IX metabolism in human bladder cancer in vitro. Photochem Photobiol 72:226–233

    Article  PubMed  CAS  Google Scholar 

  9. Jonker JW, Musters S, Vlaming ML, Plösch T, Gooijert KE, Hillebrand MJ, Rosing H, Beijnen JH, Verkade HJ, Schinkel AH (2007) Breast cancer resistance protein (Bcrp1/Abcg2) is expressed in the harderian gland and mediates transport of conjugated protoporphyrin IX. Am J Physiol Cell Physiol 292:C2204–C2212. doi:10.1152/ajpcell.00359.2006

    Article  PubMed  CAS  Google Scholar 

  10. Zhou S, Zong Y, Ney PA, Nair G, Stewart CF, Sorrentino BP (2005) Increased expression of the Abcg2 transporter during erythroid maturation plays a role in decreasing cellular protoporphyrin IX levels. Blood 105:2571–2576. doi:10.1182/blood-2004-04-1566

    Article  PubMed  CAS  Google Scholar 

  11. Asashima T, Hori S, Ohtsuki S, Tachikawa M, Watanabe M, Mukai C, Kitagaki S, Miyakoshi N, Terasaki T (2006) ATP-binding cassette transporter G2 mediates the efflux of phototoxins on the luminal membrane of retinal capillary endothelial cells. Pharm Res 23:1235–1242. doi:10.1007/s11095-006-0067-2

    Article  PubMed  CAS  Google Scholar 

  12. Robey RW, Steadman K, Polgar O, Bates SE (2005) ABCG2-mediated transport of photosensitizers: potential impact on photodynamic therapy. Cancer Biol Ther 4:187–194

    Article  PubMed  CAS  Google Scholar 

  13. Chu ES, Yow CM, Shi M, Ho RJ (2008) Effects of photoactivated 5-aminolevulinic acid hexyl ester on MDR1 over-expressing human uterine sarcoma cells. Toxicol Lett 181:7–12. doi:10.1016/j.toxlet.2008.06.860

    Article  PubMed  CAS  Google Scholar 

  14. Liu W, Baer MR, Bowman MJ, Pera P, Zheng X, Morgan J, Pandey RA, Oseroff AR (2007) The tyrosine kinase inhibitor imatinib mesylate enhances the efficacy of photodynamic therapy by inhibiting ABCG2. Clin Cancer Res 13:2463–2470. doi:10.1158/1078-0432.CCR-06-1599

    Article  PubMed  CAS  Google Scholar 

  15. Diestra JE, Scheffer GL, Català I, Maliepaard M, Schellens JH, Scheper RJ, Germà-Lluch JR, Izquierdo MA (2002) Frequent expression of the multi-drug resistance-associated protein BCRP/MXR/ABCP/ABCG2 in human tumours detected by the BXP-21 monoclonal antibody in paraffin-embedded material. J Pathol 198:213–219. doi:10.1002/path.1203

    Article  PubMed  CAS  Google Scholar 

  16. Morgan J, Jackson JD, Zheng X, Pandey SK, Pandey RK (2010) Substrate affinity of photosensitizers derived from chlorophyll-a: the ABCG2 transporter affects the phototoxic response of side population stem cell-like cancer cells to photodynamic therapy. Mol Pharm 7:1789–1804. doi:10.1021/mp100154j

    Article  CAS  Google Scholar 

  17. Sharma S, Jajoo A, Dube A (2007) 5-Aminolevulinic acid-induced protoporphyrin-IX accumulation and associated phototoxicity in macrophages and oral cancer cell lines. J Photochem Photobiol B 88:156–162. doi:10.1016/j.jphotobiol.2007.07.005

    Article  PubMed  CAS  Google Scholar 

  18. Rigby CC, Franks LM (1970) A human tissue culture cell line from a transitional cell tumour of the urinary bladder: growth, chromosone pattern and ultrastructure. Br J Cancer 24:746–754

    Article  PubMed  CAS  Google Scholar 

  19. Inoue K, Karashima T, Kamada M, Shuin T, Kurabayashi A, Furihata M, Fujita H, Utsumi K, Sasaki J (2009) Regulation of 5-aminolevulinic acid-mediated protoporphyrin IX accumulation in human urothelial carcinomas. Pathobiology 76:303–314. doi:10.1159/000245896

    Article  PubMed  CAS  Google Scholar 

  20. Hirai K, Sasahira T, Ohmori H, Fujii K, Kuniyasu H (2007) Inhibition of heme oxygenase-1 by zinc protoporphyrin IX reduces tumor growth of LL/2 lung cancer in C57BL mice. Int J Cancer 120:500–505. doi:10.1002/ijc.22287

    Article  PubMed  CAS  Google Scholar 

  21. Wyld L, Burn JL, Reed MW, Brown NJ (1997) Factors affecting aminolaevulinic acid-induced generation of protoporphyrin IX. Br J Cancer 76:705–712

    Article  PubMed  CAS  Google Scholar 

  22. Arita K, Yamamoto Y, Takehara Y, Utsumi T, Kanno T, Miyaguchi C, Akiyama J, Yoshioka T, Utsumi K (2003) Mechanisms of enhanced apoptosis in HL-60 cells by UV-irradiated n-3 and n-6 polyunsaturated fatty acids. Free Radic Biol Med 35:189–199

    Article  PubMed  CAS  Google Scholar 

  23. Liu YL, Ang SO, Weigent DA, Prchal JT, Bloomer JR (2004) Regulation of ferrochelatase gene expression by hypoxia. Life Sci 75:2035–2043. doi:10.1016/j.lfs.2004.03.027

    Article  PubMed  CAS  Google Scholar 

  24. Furukawa T, Wakabayashi K, Tamura A, Nakagawa H, Morishima Y, Osawa Y, Ishikawa T (2009) Major SNP (Q141 K) variant of human ABC transporter ABCG2 undergoes lysosomal and proteasomal degradations. Pharm Res 26:469–479. doi:10.1007/s11095-008-9752-7

    Article  PubMed  CAS  Google Scholar 

  25. Gewin L, Galloway DA (2001) E box-dependent activation of telomerase by human papillomavirus type 16 E6 does not require induction of c-myc. J Virol 75:7198–7201. doi:10.1128/JVI.75.15.7198-7201.2001

    Article  PubMed  CAS  Google Scholar 

  26. Ji Z, Yang G, Vasovic V, Cunderlikova B, Suo Z, Nesland JM, Peng Q (2006) Subcellular localization pattern of protoporphyrin IX is an important determinant for its photodynamic efficiency of human carcinoma and normal cell lines. J Photochem Photobiol B 84:213–220. doi:10.1016/j.jphotobiol.2006.03.006

    Article  PubMed  CAS  Google Scholar 

  27. Evseenko DA, Murthi P, Paxton JW, Reid G, Emerald BS, Mohankumar KM, Lobie PE, Brennecke SP, Kalionis B, Keelan JA (2007) The ABC transporter BCRP/ABCG2 is a placental survival factor, and its expression is reduced in idiopathic human fetal growth restriction. FASEB J 21:3592–3605. doi:10.1096/fj.07-8688com

    Article  PubMed  CAS  Google Scholar 

  28. Okimura Y, Fujita H, Ogino T, Inoue K, Shuin T, Yano H, Yasuda T, Inoue M, Utsumi K, Sasaki J (2007) Regulation of 5-aminolevulinic acid-dependent protoporphyrin IX accumulations in human histiocytic lymphoma U937 cells. Physiol Chem Phys Med NMR 39:69–82

    PubMed  CAS  Google Scholar 

  29. Amo T, Kawanishi N, Uchida M, Fujita H, Oyanagi E, Utsumi T, Ogino T, Inoue K, Shuin T, Utsumi K, Sasaki J (2009) Mechanism of cell death by 5-aminolevulinic acid-based photodynamic action and its enhancement by ferrochelatase inhibitors in human histiocytic lymphoma cell line U937. Cell Biochem Funct 27:503–515. doi:10.1002/cbf.1603

    Article  PubMed  CAS  Google Scholar 

  30. Gibson SL, Havens JJ, Metz L, Hilf R (2001) Is delta-aminolevulinic acid dehydratase rate limiting in heme biosynthesis following exposure of cells to delta-aminolevulinic acid? Photochem Photobiol 73:312–317

    Article  PubMed  CAS  Google Scholar 

  31. Fukuda H, Batlle AM, Riley PA (1993) Kinetics of porphyrin accumulation in cultured epithelial cells exposed to ALA. Int J Bioch 25:1407–1410

    Article  CAS  Google Scholar 

  32. Ding Y, Lin B, Huie CW (2001) Binding studies of porphyrins to human serum albumin using affinity capillary electrophoresis. Electrophoresis 22:2210–2216. doi:10.1002/1522-2683(20017)22:11<2210:AID-ELPS2210>3.0.CO;2-W

    Article  PubMed  CAS  Google Scholar 

  33. Taketani S, Kakimoto K, Ueta H, Masaki R, Furukawa T (2003) Involvement of ABC7 in the biosynthesis of heme in erythroid cells: interaction of ABC7 with ferrochelatase. Blood 101:3274–3280. doi:10.1182/blood-2002-04-1212

    Article  PubMed  CAS  Google Scholar 

  34. Susanto J, Lin YH, Chen YN, Shen CR, Yan YT, Tsai ST, Chen CH, Shen CN (2008) Porphyrin homeostasis maintained by ABCG2 regulates self-renewal of embryonic stem cells. PLoS One 3:e4023. doi:10.1371/journal.pone.0004023

    Article  PubMed  Google Scholar 

  35. Yamamoto K, Suzu S, Yoshidomi Y, Hiyoshi M, Harada H, Okada S (2007) Erythroblasts highly express the ABC transporter Bcrp1/ABCG2 but do not show the side population (SP) phenotype. Immunol Lett 114:52–58. doi:10.1016/j.imlet.2007.08.008

    Article  PubMed  CAS  Google Scholar 

  36. Kriska T, Korytowski W, Girotti AW (2002) Hyperresistance to photosensitized lipid peroxidation and apoptotic killing in 5-aminolevulinate-treated tumor cells overexpressing mitochondrial GPX4. Free Radic Biol Med 33:1389–1402

    Article  PubMed  CAS  Google Scholar 

  37. Kriska T, Korytowski W, Girotti AW (2005) Role of mitochondrial cardiolipin peroxidation in apoptotic photokilling of 5-aminolevulinate-treated tumor cells. Arch Biochem Biophys 433:435–446. doi:10.1016/j.abb.2004.09.025

    Article  PubMed  CAS  Google Scholar 

  38. Schoenfeld N, Mamet R, Nordenberg Y, Shafran M, Babushkin T, Malik Z (1994) Protoporphyrin biosynthesis in melanoma B16 cells stimulated by 5-aminolevulinic acid and chemical inducers: characterization of photodynamic inactivation. Int J Cancer 56:106–112

    Article  PubMed  CAS  Google Scholar 

  39. Desuzinges-Mandon E, Arnaud O, Martinez L, Huché F, Di Pietro A, Falson P (2010) ABCG2 transports and transfers heme to albumin through its large extracellular loop. J Biol Chem 285:33123–33133. doi:10.1074/jbc.M110.139170

    Article  PubMed  CAS  Google Scholar 

  40. Solazzo M, Fantappiè O, D’Amico M, Sassoli C, Tani A, Cipriani G, Bogani C, Formigli L, Mazzanti R (2009) Mitochondrial expression and functional activity of breast cancer resistance protein in different multiple drug-resistant cell lines. Cancer Res 69:7235–7242. doi:10.1158/0008-5472.CAN-08-4315

    Article  PubMed  CAS  Google Scholar 

  41. Frank J, Lornejad-Schäfer MR, Schöffl H, Flaccus A, Lambert C, Biesalski HK (2007) Inhibition of heme oxygenase-1 increases responsiveness of melanoma cells to ALA-based photodynamic therapy. Int J Oncol 31:1539–1545

    PubMed  CAS  Google Scholar 

Download references

Acknowledgments

The authors are grateful to Dr. Shigeru Taketani for providing ferrochelatase antibody. The authors are also grateful to Dr. Tohru Tanaka for the support of this investigation. This work was supported in part by grants from the Ministry of Education, Science and Culture of Japan.

Conflict of interest

None.

Author information

Authors and Affiliations

  1. Department of Pathology and Experimental Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, 2-5-1 Shikata, Okayama, 700-8558, Japan

    Tetsuya Ogino

  2. Cell Chemistry, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, 700-8558, Japan

    Hirotsugu Kobuchi

  3. Cytology and Histology, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, 700-8558, Japan

    Kazuaki Munetomo, Hirofumi Fujita, Masanao Yamamoto, Junzo Sasaki & Kozo Utsumi

  4. Department of Biological Chemistry, Faculty of Agriculture, Yamaguchi University, Yamaguchi, 753-8515, Japan

    Toshihiko Utsumi

  5. Department of Urology, Kochi University Medical School, Nankoku, 783-8505, Kochi, Japan

    Keiji Inoue & Taro Shuin

  6. Department of Biochemistry and Molecular Pathology, Osaka City University Medical School, Osaka, 545-8585, Japan

    Masayasu Inoue

Authors
  1. Tetsuya Ogino
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Hirotsugu Kobuchi
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kazuaki Munetomo
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hirofumi Fujita
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Masanao Yamamoto
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Toshihiko Utsumi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Keiji Inoue
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Taro Shuin
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Junzo Sasaki
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Masayasu Inoue
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Kozo Utsumi
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Tetsuya Ogino.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ogino, T., Kobuchi, H., Munetomo, K. et al. Serum-dependent export of protoporphyrin IX by ATP-binding cassette transporter G2 in T24 cells. Mol Cell Biochem 358, 297–307 (2011). https://doi.org/10.1007/s11010-011-0980-5

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11010-011-0980-5

Keywords

Search

Navigation