Send Orders of Reprints at reprints@benthamscience.net 932 Anti-Cancer Agents in Medicinal Chemistry, 2013, 13, 932-951 Structural Effects of TiO 2 Nanoparticles and Doxorubicin on DNA and their Antiproliferative Roles in T47D and MCF7 Cells Azadeh Hekmat 1 , Ali Akbar Saboury 1, * , Adeleh Divsalar 2 and Arefeh Seyedarabi 1,3 1 Institute of Biochemistry and Biophysics, Tehran University, Tehran, Iran; 2 Department of Biological Sciences, Tarbiat Moallem University, Tehran, Iran; 3 Institute of Structural and Molecular Biology, University College London, London, United Kingdom Abstract: The structural changes in DNA caused by the combined effects of TiO2 nanoparticles (TiO2 NPs) and doxorubicin (DOX) were investigated along with their corresponding inhibitory roles in the growth of T47D and MCF7 cells. The UV-visible titration studies showed that DOX+ TiO2 NPs could form a novel complex with DNA. The data also reveal that the TiO2–DOX complex forms through a 1:4 stoichiometric ratio in solution. The values of binding constants reveal that DOX+TiO2 NPs interact more strongly with DNA as compared to TiO2 NPs or DOX alone. CD data show that DOX+TiO2 NPs can noticeably cause disturbance on DNA structure compared to TiO2 NPs or DOX alone, considering that DNA is relatively thermally stable in the condition used. The anticancer property of 0.3 M DOX+ 60 M TiO2 NPs and 0.4 M DOX+ 670 M TiO2 NPs by MTT assay and DAPI stain demonstrates that this combination can tremendously diminish proliferation of T47D and MCF7cells compared to DOX or TiO2 NPs alone. The UV–Vis absorption spectroscopy, flow cytometry and fluorescence microscopy experiments show much more enhancement of DOX uptake through the use of TiO2 NPs. These results reveal that DOX+TiO2 NPs could proffer a novel strategy for the development of promising and efficient chemotherapy agents. Keywords: Anthracycline, Anti-Cancer drug uptake, Breast cancer cell line T47D, Breast cancer cell line MCF7, Chemotherapeutic agent, Circular dichroism (CD), DAPI stain, DNA structure, Doxorubicin (Adriamycin), Drug combination, Flow cytometry, Fluorescence spectroscopy, Fluorescence microscopy, Human adult stem cell, MTT assay, Titanium dioxide nanoparticles (TiO 2 NPs), UV-Visible spectroscopy. INTRODUCTION For almost half a century, nanoparticles (NPs) have been attracting a great deal of attention by scientists owing to their distinctive chemical, physical, biological as well as functional properties [1]. The fast-growing progresses in the field of nanotechnology by physical methods including optical spectroscopy haveprovided a great opportunity to obtain new insights into the interaction mechanisms of biomolecules with various NPs. It is worth noting that exploring the behavior of eukaryotic cells as well as prokaryotic cells after exposure to NPs has attracted tremendous attention in the field of nanomedicine [2]. Among the semiconductor NPs, titanium dioxide nanoparticles (TiO 2 NPs) have been utilized most widely in the nanotechnology industry due to their chemical inertness, strong oxidizing properties, photoreactivity and high stability [3]. It is important to mention that titanium is a very reactive metal, which is readily oxidized via diverse media. The oxidation kinetics is extremely fast in, which initial events occurr on a time scale of few nanoseconds. Owing to the high reactivity of titanium and short time scales involved in the oxidation, titanium is almost always covered by an oxidized layer. In almost cases, the oxidized surface layer consists mainly of the most stable TiO 2 . In contrast to titanium, TiO 2 is a stable compound, which is resistant to chemical attack from most substances. The excellent chemical and corrosion resistance of titanium is to a large extent due to chemical stability of its surface oxide. The latter must not, however, be confused with chemical inertness. On the contrary, TiO 2 surfaces will react with aqueous solutions and they will irreversibly adsorb and dissociate organic molecules from air. It is also well known in the surface science community that TiO 2 surfaces act as catalysts for a number of chemical reactions. Thus, TiO 2 is stable but chemically active [4]. *Address correspondence to this author at the Institute of Biochemistry and Biophysics, Tehran University, Tehran, Iran; Tel: +98-21-66956984; Fax: +98-21-66404680; E-mail: saboury@ut.ac.ir TiO 2 NPs exists in three different crystallographic structures: rutile, anatase and brookite, in which the anatase phase exhibits a better biocompatibility in comparison with rutile. The cytotoxicity induced in human colon cells, bronchial epithelial cells and human hepatoma cells was observed after treatment with TiO 2 NPs [5, 6]. Current studies on cytotoxicity activity of TiO 2 NPs in human cells demonstrated that TiO 2 NPs could induce lipid peroxidation, lysosomal membrane destabilization, release of cytochrome c from mitochondria, reactive oxygen species (ROS) generation and also DNA damage [7-10]. The anthracycline antibiotic doxorubicin (DOX; former generic name, Adriamycin); which is isolated from the fungus Streptomyces peucetius; can remarkably hinder the growth of cancer cells and has been utilized since 1970s for the treatment of a wide range of human tumors like leukemia, ovarian cancer and late stage breast cancer [11]. The molecular structure of DOX is given in Scheme 1. Numerous different mechanisms have been suggested for the function of DOX against cancer cells, namely, the inhibition of nucleic acid and protein synthesis, membrane interaction and the production of ROS [11-13]. Unfortunately, DOX causes a number of serious adverse effects including myelosuppression, nausea and cumulative cardiotoxicity [14]. Owing to the harmful and strong adverse reactions in patients and the red color of DOX, this drug has acquired the nickname red death or red devil [15]. Moreover, multidrug resistance (MDR) of cancer cells to DOX remains the most important barrier to a successful cancer treatment [16]. The continuous frequency of resistance to chemotherapeutic agents, their low efficacy towards tumor cells and their serious side effects in cancer patients have placed a demand on the scientific community to constantly design novel chemotherapeutics drugs. Countless empirical and theoretical methods have been sought to recover the affinity and specificity of anticancer drugs towards DNA molecules as well as to diminish their hazardous side effects. Some reports emphasize that combining more than one anticancer drug can defeat adverse side effects correlated with high doses of a single drug [17, 18]. 1875-5992/13 $58.00+.00 © 2013 Bentham Science Publishers