Abstract
Background
Human multiple myeloma (MM) remains an incurable hematological malignancy. We have reported that β-lapachone, a pure compound derived from a plant, can induce cell death in a variety of human carcinoma cells, including ovary, colon, lung, prostate, pancreas, and breast, suggesting a wide spectrum of anticancer activity.
Materials and Methods
We first studied anti-survival effects of β-lapachone in human MM cells by colony formation assay. To determine whether the differential inhibition of colony formation occurs through antiproliferative activity, we performed MTT assays. The cytotoxicity of β-lapachone on human peripheral blood mononuclear cells was also measured by MTT assay. To determine whether the cell death induced by β-lapachone occurs through necrosis or apoptosis, we used the propidium iodide staining procedure to determine the sub-G1 fraction, Annexin-V staining for externalization of phosphatidylserine, and fragmentation of cellular genomic DNA subjected to gel electrophoresis. To investigate the mechanism of anti-MM activity, we examined Bcl-2 expression, cytochrome C release, and poly (ADP ribose) polymerase cleavage by Western blot assay.
Results
We found that β-lapachone (less than 4 µM) inhibits cell survival and proliferation by triggering cell death with characteristics of apoptosis in ARH-77, HS Sultan, and MM.1S cell lines, in freshly derived patient MM cells (MM.As), MM cell lines resistant to dexamethasone (MM.1R), doxorubicin (DOX.40), mitoxantrone (MR.20), and mephalan (LR5). Importantly, after treatment with β-lapachone, we observed no apoptosis in peripheral blood mononuclear cells in either quiescent or proliferative states, freshly isolated from healthy donors. In β-lapachone treated ARH-77, cytochrome C was released from mitochondria to cytosol, and poly (ADP ribose) polymerase was cleaved, signature events of apoptosis. Finally, the apoptosis induced by β-lapachone in MM cells was not blocked by either interleukin-6 or Bcl-2, which confer multidrug resistance in MM.
Conclusions
Our results suggest potential therapeutic application of β-lapachone against MM, particularly to overcome drug resistance in relapsed patients.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Treon SP, Teoh G, Urashima M, Ogata A, Chauhan D, Webb J, Anderson K. (1998) Antiestrogens induce apoptosis of multiple myeloma cells. Blood 92: 1749–1757.
Anderson K, Kyle RA, Berenson JR, Dalton WS.(1999) Recent Advances in the Biology and Treatment of Multiple Myeloma. Annual Meeting Report 1999.
Hooker SC. (1936) Lomatiol. Part II. Its occurence, constitution, relation to and conversion into lapachol. Also a synthesis of lapachol J. Am. Chem. Soc. 58: 1181–1190.
Goncalves de Lima O, D’Albuquerque IL, Goncalves de Lima C, Dalla Maia MH. (1962) Substancias antimicrobianas de plantas superiores. Rev. Inst. Antibiot. Univ. Recife. 4: 3–17.
Li CJ, Averboukh L, Pardee AB. (1993) Betalapachone, a novel DNA topoisomerase I inhibitor with a mode of action different from camptothecin. J. Biol. Chem. 268: 22463–22468.
Goncalves AM, Vasconcellos ME, Docampo R, Cruz NS, Souza WR, Leon W. (1980) Evaluation of the toxicity of 3-allyl-β-lapachone against trypanosoma cruzi bloodstream forms. Mol. Biochem. Parasitology. 1: 167–176.
Schaffner-Sabba K, Schmidt-Ruppin KH, Wehrli W, Schuerch AR, Wasley JWF. (1984) β-Lapachone: synthesis of derivatives and activities in tumor models. J. Med. Chem. 27: 990–994.
Li CJ, Zhang LJ, Dezube BJ, Crumpacker CS, Pardee AB. (1993) Three inhibitors of type I human immunodeficiency virus long terminal repeat-directed gene expression and virus replication. Proc. Natl. Acad. Sci. USA. 90: 1839–1842.
Schuerch AR, Wehrli W. (1978) β-Lapachone, an inhibitor of oncornavirus reverse transcriptase and eukaryotic DNA polymerase-α. Eur. J. Biochem. 84: 197–205.
Boorstein RJ, Pardee AB. (1984) β-Lapachone greatly enhances MMS lethality to human fibroblasts. Biochem. Biophys. Res. Commun. 118: 828–834.
Boothman DA, Trask DK, Pardee AB. (1989) Inhibition of potentially lethal DNA damage repair in human tumor cells by β-lapachone, an activator of topoisomerase I. Cancer Res. 49: 605–612.
Li CJ, Averboukh L, Pardee AB. (1993) β-Lapachone, a novel DNA topoisomerase I inhibitor with a mode of action different from camptothecin. J. Biol. Chem. 268: 22463–22468.
Li YZ, Li CJ, Pinto AV, Pardee AB. (1999) Release of mitochondrial cytochrome C in both apoptosis and necrosis induced by β-lapachone in human carcinoma cells. Molec. Med. 5: 232–239.
Tu Y, Xu FH, Liu J, Vescio R, Berenson J, Fady C, Lichtenstein A. (1996) Upregulated expression of BCL-2 in multiple myeloma cells induced by exposure to doxorubicin, etoposide, and hydrogen peroxide. Blood 88: 1805–1812.
Bloem A, Lockhorst H. (1999) Bcl-2 antisense therapy in multiple myeloma. Pathol. Biol. (Paris) 47: 216–220.
Anderson K. (1999) Advances in the biology of multiple myeloma: therapeutic applications. Sem. Oncol. 26(suppl 13): 10–22.
Mosmann T. (1983) Rapid colorimertic assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Meth. 65: 55.
Anderson K. (1999) Introduction. Sem. Oncol. 26: 1.
Li CJ, Friedman DJ, Wang C, Metelev V, Pardee AB. (1995) Induction of apoptosis in uninfected lymphocytes by HIV-1 tat protein. Science 268: 429–431.
Li CJ, Li YZ, Pinto AV, Pardee A. (1999) Potent inhibition of tumor survival in vivo by β-lapachone plus taxol: combining drugs imposes different artificial checkpoints. Proc. Natl. Acad. Sci. USA 96: 13369–13374.
Zhang G, Gurtu V, Kain SR, Yan G. (1997) Early detection of apoptosis using a fluorescent conjugate of annexin V. BioTechniques 23: 525–531.
Li CJ, Wang C, Pardee AB. (1995) Induction of apoptosis by β-lapachone in human prostate cancer cells. Cancer Res. 55: 3712–3715.
Rita S, Michele R, Patrizia O, Alessandra B, Sara B, Enrico G. (1994) Inhibition of protein kinase C in PHA-activated PBMC treated with anti-HLA class I monoclonal antibody is associated to decreased proliferation and expression of cell cycle related genes. Biochem. Mol. Biol. Int. 32: 105–114.
Case DC, Jr., Lee, DJ III, Clarkson BD. (1977) Improved survival times in multiple myeloma treated with melphalan, prednisone, cyclophosphamide, vincristine and BCNU: M-2 protocol. Am. J. Med. 63: 897–903.
Otsuki T, Yamada O, Kurebayashi J, et al. (2000) Estrogen receptors in human myeloma cells. Cancer Res. 60: 1434–1441.
Acknowledgments
We thank Dr. Kenneth Anderson for providing MM cells and for critically reading the manuscript. This work was supported in part by Grant RO1 CA61253 from the National Institutes of Health.
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Li, Y., Li, C.J., Yu, D. et al. Potent Induction of Apoptosis by β-Lapachone in Human Multiple Myeloma Cell Lines and Patient Cells. Mol Med 6, 1008–1015 (2000). https://doi.org/10.1007/BF03402052
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/BF03402052