The cancer growth suppressing gene mda-7 induces apoptosis selectively in human melanoma cells Irina V Lebedeva 1,2,5 , Zao-zhung Su 1,2,5 , Yonmee Chang 1,2 , Shinichi Kitada 3 , John C Reed 3 and Paul B Fisher* ,1,2,4 1 Department of Pathology, Herbert Irving Comprehensive Cancer Center, Columbia University, College of Physicians and Surgeons, 630 West 168th Street, New York, NY 10032, USA; 2 Department of Urology, Herbert Irving Comprehensive Cancer Center, Columbia University, College of Physicians and Surgeons, 630 West 168th Street, New York, NY 10032, USA; 3 The Burnham Institute, 10901 North Torrey Pines Road, La Jolla, California, CA 92037, USA; 4 Department of Neurosurgery, Herbert Irving Comprehensive Cancer Center, Columbia University, College of Physicians and Surgeons, 630 West 168th Street, New York, NY 10032, USA Human melanoma cells growth arrest irreversibly, lose tumorigenic potential and terminally dierentiate after treatment with a combination of fibroblast interferon (IFN-b) and the protein kinase C activator mezerein (MEZ). Applying subtraction hybridization to this model dierentiation system permitted cloning of melanoma dierentiation associated gene-7, mda-7. Expression of mda-7 inversely correlates with melanoma development and progression, with elevated expression in normal melanocytes and nevi and increasingly reduced expres- sion in radial growth phase, vertical growth phase and metastatic melanoma. When expressed by means of a replication incompetent adenovirus (Ad.mda-7) growth of melanoma, but not normal early passage or immortal human melanocytes, is dramatically suppressed and cells undergo programmed cell death (apoptosis). Infection of metastatic melanoma cells with Ad.mda-7 results in an increase in cells in the G 2 /M phase of the cell cycle and changes in the ratio of pro-apoptotic (BAX, BAK) to anti-apoptotic (BCL-2, BCL-XL) proteins. Ad.mda-7 infection results in a temporal increase in mda-7 mRNA and intracellular MDA-7 protein in most of the melanocyte/melanoma cell lines and secretion of MDA- 7 protein is readily detected following Ad.mda-7 infection of both melanocytes and melanoma cells. The present studies document a dierential response of melanocytes versus melanoma cells to ectopic expression of mda-7 and support future applications of mda-7 for the gene- based therapy of metastatic melanoma. Oncogene (2002) 21, 708 – 718. DOI: 10.1038/sj/onc/ 1205116 Keywords: melanoma dierentiation associated gene-7; tumor suppressor; cell cycle; cancer specific cytokine; gene therapy; BCL-family Introduction A useful experimental model for defining the genetic and biochemical changes underlying tumor progression is metastatic human melanoma (Herlyn, 1990; Lu and Kerbel, 1994; Leszczyniecka et al., 2001). Of the numerous types of cancer developing in North American populations, melanoma is increasing at the fastest rate and it is estimated that as many as one in 75 currently born Caucasian children may eventually develop superficial spreading type melanoma (Clark, 1991; Armstrong and Kricker, 1994). Although melanoma is curable at early stages, surgical and chemotherapeutic interventions are virtually ineective in preventing metastatic disease and death in patients with advanced stages of malignant melanoma. These findings underscore the need for improved therapeutic approaches to more ecaciously treat patients with metastatic melanoma. Development of malignant melanoma in humans, with the exception of nodular type melanoma, may involve a series of sequential alterations in the evolving tumor cells (Herlyn, 1990; Clark, 1991; Armstrong and Kricker, 1994; Lu and Kerbel, 1994; Leszczyniecka et al., 2001). Although several models of progression are possible, a proposed scheme for cutaneous melanoma development consists of the conversion of a normal melanocyte into a common acquired melanocytic nevus (mole), followed by the development of a dysplastic nevus, a radial growth phase (RGP) primary melanoma, a vertical growth phase (VGP) primary melanoma and ultimately a metastatic melanoma (Herlyn, 1990; Clark, 1991; Armstrong and Kricker, 1994; Lu and Kerbel, 1994; Leszczyniecka et al., 2001). Although primary melanoma is readily treatable, even during the VGP if the lesion is 40.76-mm thick, currently employed techniques are minimally eective (520% survival) in preventing metastatic spread and morbidity in patients with VGP lesions of 44.0-mm thickness. Cancer progression has been extensively studied in the context of human melanocytes, nevi, primary melanomas and metastatic melanomas (Herlyn, 1990; Clark, 1991; Armstrong and Kricker, 1994; Lu and Kerbel, 1994; Oncogene (2002) 21, 708 – 718 ª 2002 Nature Publishing Group All rights reserved 0950 – 9232/02 $25.00 www.nature.com/onc *Correspondence: PB Fisher, Departments of Pathology and Urology, Columbia University, College of Physicians and Surgeons, BB-1501, 630 West 168th Street, New York, NY 10032, USA; E-mail: pbfl@columbia.edu 5 These authors contributed equally to this manuscript Received 16 August 2001; revised 22 October 2001; accepted 30 October 2001