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Research Articles: Therapeutics, Targets, and Development

The role of autophagy in the death of L1210 leukemia cells initiated by the new antitumor agents, XK469 and SH80

¹ Department of Pharmacology, Wayne State University School of Medicine; ² Institute of Environmental Health Sciences, Wayne State University; and ³ Department of Internal Medicine, Division of Hematology and Oncology, Wayne State University School of Medicine and Barbara Ann Karmanos Cancer Institute, Detroit, Michigan

Requests for reprints: David Kessel, Department of Pharmacology, Wayne State University School of Medicine, Detroit, MI 48201. Phone: 313-577-1766; Fax: 313-577-6739. E-mail: dhkessel{at}med.wayne.edu

Abstract

The phenoxypropionic acid derivative 2-{4-[(7-chloro-2-quinoxalinyl)oxy]phenoxy}propionic acid (XK469) and an analogue termed 2-{4-[(7-bromo-2-quinalinyl)oxy]phenoxy}propionic acid (SH80) can eradicate malignant cell types resistant to many common antitumor agents. Colony formation assays indicated that a 24 h exposure of L1210 cells to XK469 or SH80 inhibited clonogenic growth with CI₉₀ values of 10 and 13 µmol/L, respectively. This effect was associated with G₂-M arrest and the absence of any detectable markers of apoptosis (i.e., plasma membrane blebbing, procaspase 3 activation, loss of mitochondrial membrane potential, and formation of condensed chromatin). Drug-treated cells increased in size and eventually exhibited the characteristics of autophagy (i.e., appearance of autophagosomes and conversion of microtubule-associated protein light chain 3-I to 3-II). The absence of apoptosis was not related to an inhibition of the apoptotic program. Cultures treated with XK469 or SH80 readily underwent apoptosis upon exposure to the Bcl-2/Bcl-x_L antagonist ethyl 2-amino-6-bromo-4-(1-cyano-2-ethoxy-2-oxoethyl)-4H-chromene-3-carboxylate. Continued incubation of drug-treated cells led to a reciprocal loss of large autophagic cells and the appearance of smaller cells that could not be stained with Höechst dye HO33342, had a chaotic morphology, were trypan blue–permeable, and lacked mitochondrial membrane potential. L1210 cells cotreated with the phosphatidylinositol-3-kinase inhibitor wortmannin, or having reduced Atg7 protein content, underwent G₂-M arrest, but not autophagy, following XK469 treatment. Hence, the therapeutic actions of XK469/SH80 with L1210 cultures reflect both the initiation of a cell cycle arrest as well as the initiation of autophagy. [Mol Cancer Ther 2007;6(1):370–9]

Grant support: National Cancer Institute grants CA-082341 (J.P. Horwitz and D. Kessel) and CA-23378 (D. Kessel and J. Reiners). Additional support from the Imaging and Cytometry Facility Core supported by NIEHS grant P30 ES06639 and the Jack and Miriam Schenkman Cancer Research Fund (J.P. Horwitz).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

⁴ D. Kessel and J.J. Reiners, Jr., unpublished data.

⁵ J.J. Reiners, Jr., unpublished data.

Received 9/26/06; revised 10/30/06; accepted 11/21/06.