Casiopeı ´na IIgly induced cytotoxicity to HeLa cells depletes the levels of reduced glutathione and is prevented by dimethyl sulfoxide Radame ´s Alemo ´ n-Medina a , Jose ´ Luis Mun ˜ oz-Sa ´nchez b , Lena Ruiz-Azuara a,1, * , Isabel Gracia-Mora a a Departamento de Quı ´mica Inorga ´ nica y Nuclear, Facultad de Quı ´mica, Universidad Nacional Auto ´noma de Me ´xico (UNAM), Me ´xico D.F. CP 04510, Mexico b Departamento de Bioquı ´mica, Escuela Nacional de Ciencias Biolo ´ gicas, Instituto Polite ´cnico Nacional (IPN), Prolongacio ´ n de Carpio s/n, Col. Casco de Santo Toma ´s, Me ´xico D.F. CP 11340, Mexico Received 14 September 2007; accepted 12 November 2007 Available online 21 November 2007 Abstract The newly synthesized copper coordination compound Casiopeı ´na IIgly (Cas IIgly) is a promising alternative drug in the treatment of cancer, since it has shown cytotoxicity and genotoxicity in different tumour models. Given its enhanced effects after ascorbic acid-med- iated copper reduction, Cas IIgly’s activity is thought to be related to oxidative damage. In the present work, oxidized Cas IIgly failed to induce cytosolic oxidative damage in HeLa cells (only 0.9% of the cell population), and in 2.3% of the treated cells when previously reduced, as evaluated through the oxidation of dihydrorhodamine 123 (DHR 123). However, it showed cytotoxicity, since HeLa cells treated with 10–80 lg/mL Cas IIgly proliferated only at 30% of their normal rate, and at 15% when treated with reduced Cas IIgly. This cytotoxicity is strongly abolished in the presence of the hydroxyl scavenger dimethyl sulfoxide. The decrease, from 3994 to 530 nano- grams of reduced glutathione (GSH) per million cells after treatment with 80 lg/mL Casiopeı ´na IIgly, indicates that this drug causes the expenditure of this naturally occurring antioxidant. These results altogether suggest that, albeit Cas IIgly induced cytotoxicity is not related to cytosolic DHR 123 oxidation, it may be related to oxidative damage. Ó 2007 Elsevier Ltd. All rights reserved. Keywords: Casiopeı ´na IIgly; Copper; Oxidative damage; Dihydrorhodamine 123 (DHR 123); Reduced glutathione (GSH) 1. Introduction The novel drug Casiopeı ´na IIgly (Cas IIgly) (Fig. 1) is a copper (Cu(II)) chelate with dimethyl phenanthroline and the aminoacid glycine as bidentate ligands (Ruiz-Ramı ´rez et al., 1993a,b; Gracia-Mora et al., 2001). Its promising use as a chemotherapeutic agent in the treatment of cancer arises from various studies conducted in vitro and in vivo, in which its ability to reduce the size and volume of cultured and implanted tumours (De Vizcaya et al., 2000; Gracia- Mora et al., 2001), its low toxicity to healthy tissues and, furthermore, its capacity to induce apoptosis in glioma C6 cells (Trejo-Solı ´s et al., 2005) have been shown. Regarding Cas IIgly mode of action, it has been demon- strated that this compound interferes both the electron transfer chain and oxidative phosphorylation in mitochon- dria isolated from cardiac muscle and glioma cells (Marı ´n- Herna ´ndez et al., 2003; Trejo-Solı ´s et al., 2005), and its ability to promote DNA fragmentation and apoptosis is as well documented previously (Rivero-Mu ¨ ller et al., 1998). Cas IIgly failed to induce lipid peroxidation to plasma membrane in HeLa cells and in human lympho- cytes (Alemo ´n-Medina et al., 2007), but produced DNA damage and cell death, which were promoted after the 0887-2333/$ - see front matter Ó 2007 Elsevier Ltd. All rights reserved. doi:10.1016/j.tiv.2007.11.011 * Corresponding author. Tel./fax: +52 5 5556223529. E-mail addresses: ranapez@hotmail.com (R. Alemo ´ n-Medina), pcervan@hotmail.com (J.L. Mun ˜ oz-Sa ´nchez), ruizazuara@gmail.com (L. Ruiz-Azuara), isabelg@servidor.unam.mx (I. Gracia-Mora). 1 Previously as Lena Ruiz-Ramı ´rez. www.elsevier.com/locate/toxinvit Available online at www.sciencedirect.com Toxicology in Vitro 22 (2008) 710–715